Fumaric acid fermentation with immobilised Rhizopus oryzae : Quantifying time-dependent variations in catabolic flux

Naude, Andre; Nicol, Willie

doi:10.1016/j.procbio.2017.02.027

Cited by 21 publications

(41 citation statements)

References 23 publications

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“…Succinic acid (C 4 H 6 O 4 ) was identified as the minor by-product; its presence could be due to the release of this compound from the cytoplasm and from mitochondria, as a major component in the TCA cycle. No ethanol was present in this occasion, though it is observed in other works as a by-product that is produced during biomass growth and disappears in the stationary phase [17,22]. Apart from this, results are very similar to those obtained in other studies [8].…”

Section: Kinetic Modellingsupporting

confidence: 90%

“…As in other studies [8,17,22], the presence of some by-products have been detected. The major by-product was identified as malic acid (C 4 H 6 O 5 ), as malic and fumaric acid concentrations are very similar [8].…”

Section: Kinetic Modellingsupporting

confidence: 84%

“…Just considering food wastes, which are not as plentiful as Most studies on fumaric acid of biotechnological origin focus on the development of an effective fermentative process for fumaric acid production at an industrial level. However, although kinetic models are of great interest to simulate and design industrial processes, during their scale-up and final production control, in the case of fumaric acid, there is, to the best of our knowledge, only a kinetic model study focused on immobilized Rhizopus oryzae, but none for the more common free or suspended fungus [22]. The kinetic Monod model proposed in this study is employed to determine yields of fumaric acid and by-products by understanding the evolution of the system through the different catabolic carbon fluxes proposed (fumaric acid production, respiration and ethanol production).…”

Section: Introductionmentioning

confidence: 99%

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Kinetic Modelling of the Coproduction Process of Fumaric and Malic Acids by Rhizopus arrhizus NRRL 1526

et al. 2020

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The production of organic acids by biotechnological processes has experienced a notable impulse with the advent of first and second generation biorefineries and the need of searching for renewable and sustainable feedstock, such as biomass. Fumaric acid is a promising biomonomer for polyamide production and a well-known acidulant and preservative in food and feed industries. Malic acid is a well-known food acidulant with a high market share. The biotechnological Fumaric and Malic acid production via fungi of the Rhizopus genus is being explored nowadays as a process for the valorization of food and food-related waste to obtain food ingredients and key platform chemicals of the so-called biochemical biorefinery. In this work, a preliminary study is performed to find reproducible conditions for the production of the acids by Rhizopus arrhizus NRRL 1526 by controlling fungi morphology and inoculum conditions. Afterwards, several production runs are performed to obtain biomass, glucose, and acid concentration data at different processing time values. Finally, an unstructured, unsegregated model including a logistic-type equation for biomass and potential-type equations for the substrate and the products is fitted to experimental data. We find that the production of the organic acids is mainly non-associated with fungal growth.

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Section: Kinetic Modellingsupporting

confidence: 90%

Section: Kinetic Modellingsupporting

confidence: 84%

Section: Introductionmentioning

confidence: 99%

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Kinetic Modelling of the Coproduction Process of Fumaric and Malic Acids by Rhizopus arrhizus NRRL 1526

et al. 2020

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“…Rhizopus oryzae (ATCC 20344) was used for all fermentations and was prepared as described by Naude and Nicol [27]. The spore inoculum used for batch growth fermentations had a spore concentration of 8 × 10 6 mL −1 of which 10 mL were aseptically injected into the reactor through a silicon septum.…”

Section: Microorganism and Culture Conditionsmentioning

confidence: 99%

“…In this study, an immobilised form of R. oryzae is used to investigate the organism's Crabtree characteristics. We have successfully employed the immobilised process in previous studies [24,26,27] and have shown that biomass-medium separation and process up-scaling are considerably simplified when compared to systems with suspended fungal pellets. The initial experimental work explores the effect of oxygen availability in the medium on ethanol formation, while subsequent experiments investigate the effect of glucose limitation on the growth and fumarate production (nitrogen limited) response in order to test the postulated response depicted in Fig.…”

Section: Introductionmentioning

confidence: 99%

Fumarate production with Rhizopus oryzae: utilising the Crabtree effect to minimise ethanol by-product formation

Swart

Roux

Naude

et al. 2020

Biotechnol Biofuels

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Background: The four-carbon dicarboxylic acids of the tricarboxylic acid cycle (malate, fumarate and succinate) remain promising bio-based alternatives to various precursor chemicals derived from fossil-based feed stocks. The double carbon bond in fumarate, in addition to the two terminal carboxylic groups, opens up an array of downstream reaction possibilities, where replacement options for petrochemical derived maleic anhydride are worth mentioning. To date the most promising organism for producing fumarate is Rhizopus oryzae (ATCC 20344, also referred to as Rhizopus delemar) that naturally excretes fumarate under nitrogen-limited conditions. Fumarate excretion in R. oryzae is always associated with the co-excretion of ethanol, an unwanted metabolic product from the fermentation. Attempts to eliminate ethanol production classically focus on enhanced oxygen availability within the mycelium matrix. In this study our immobilised R. oryzae process was employed to investigate and utilise the Crabtree characteristics of the organism in order to establish the limits of ethanol by-product formation under growth and non-growth conditions. Results: All fermentations were performed with either nitrogen excess (growth phase) or nitrogen limitation (production phase) where medium replacements were done between the growth and the production phase. Initial experiments employed excess glucose for both growth and production, while the oxygen partial pressure was varied between a dissolved oxygen of 18.4% and 85%. Ethanol was formed during both growth and production phases and the oxygen partial pressure had zero influence on the response. Results clearly indicated that possible anaerobic zones within the mycelium were not responsible for ethanol formation, hinting that ethanol is formed under fully aerobic conditions as a metabolic overflow product. For Crabtree-positive organisms like Saccharomyces cerevisiae ethanol overflow is manipulated by controlling the glucose input to the fermentation. The same strategy was employed for R. oryzae for both growth and production fermentations. It was shown that all ethanol can be eliminated during growth for a glucose addition rate of 0.07 g L −1 h −1. The production phase behaved in a similar manner, where glucose addition of 0.197 g L −1 h −1 resulted in fumarate production of 0.150 g L −1 h −1 and a yield of 0.802 g g −1 fumarate on glucose. Further investigation into the effect of glucose addition revealed that ethanol overflow commences at a glucose addition rate of 0.395 g g −1 h −1 on biomass, while the maximum glucose uptake rate was established to be between 0.426 and 0.533 g g −1 h −1. Conclusions: The results conclusively prove that R. oryzae is a Crabtree-positive organism and that the characteristic can be utilised to completely discard ethanol by-product formation. A state referred to as "homofumarate production" was illustrated, where all carbon input exits the cell as either fumarate or respiratory CO 2. The highest biomass-based

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Malic acid production by Aspergillus oryzae: the immobilized fungal fermentation route

Brink

Geyer‐Johnson

Swart

et al. 2022

Biofuels Bioprod Bioref

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It has been proposed that the microbial bioproduction of chemical precursors could reduce societal overreliance on the petrochemical industry. However, economic implementation remains elusive due to a lack of understanding of the mechanisms involved. The current study evaluated malic acid production using a novel immobilized Aspergillus oryzae fungal bed bioreactor. The experimental procedures involved two different nitrogen sources (urea and (NH4)2SO4), two‐phase growth/production and simultaneous single‐step growth/production runs, the presence and absence of added CO2, the presence and absence of pH control, and the use of glucose and dl‐malate as the carbon sources. The experimental malic acid titers were remarkably poor (between 1.7 and 3.3 g.L−1) in comparison with CaCO3 buffered malic acid titers from literature (concentrations in excess of 100 g.L−1 have been reported reported). In contrast, citric acid was observed to be the main metabolite. The dl‐malate carbon source was consumed with concomitant growth, indicating A. oryzae′s ability to utilize both d and l enantiomers of malic acid. Detailed metabolic flux analyses predicted partial consumption of urea during growth, while (NH4)2SO4 was fully utilized. During the urea runs, significant amounts of glycogen were accumulated in the biomass – which were utilized as substrate during the production run. Nitrogen limitations induced significant lipid production, corresponding to citric acid accumulation in the media; the mechanism for fungal lipid biosynthesis involves citric acid excretion at the expense of malic acid. These results indicate a possible calcareous trigger for malic acid production as opposed to the traditionally accepted nitrogen starvation mechanism – nitrogen starvation resulted in lipid accumulation with corresponding malate loss. © 2022 The Authors. Biofuels, Bioproducts and Biorefining published by Society of Industrial Chemistry and John Wiley & Sons Ltd.

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Fumaric acid fermentation with immobilised Rhizopus oryzae : Quantifying time-dependent variations in catabolic flux

Cited by 21 publications

References 23 publications

Kinetic Modelling of the Coproduction Process of Fumaric and Malic Acids by Rhizopus arrhizus NRRL 1526

Kinetic Modelling of the Coproduction Process of Fumaric and Malic Acids by Rhizopus arrhizus NRRL 1526

Fumarate production with Rhizopus oryzae: utilising the Crabtree effect to minimise ethanol by-product formation

Malic acid production by Aspergillus oryzae: the immobilized fungal fermentation route

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