Citric Acid Biosynthesis Research Articles

Enhanced Citric Acid Production through Aspergillus niger: Insights from Fermentation Studies Using Sugarcane Molasses.

The production of citric acid, a vital agricultural commodity utilized across various industries such as food, beverages, pharmaceuticals, agriculture, detergents, and cosmetics, predominantly relies on microbial fermentation, with Aspergillus niger accounting for approximately 90% of global production. In this study, we aimed to optimize the key factors influencing citric acid production, with a focus on strains, fermentation techniques, and carbon sources, particularly sugarcane molasses. A. niger, sourced from the Botany department/Biotechnology laboratories at Govt. College of Science, Lahore, was employed for citric acid production. The process involved inoculum preparation through spore collection from 3 to 5 days of cultured PDA slants. The fermentation medium, comprising cane molasses with a 15% sugar concentration, was meticulously prepared and optimized for various factors, including magnesium sulfate, potassium ferrocyanide, time of addition of potassium ferrocyanide, ammonium oxalate, and calcium chloride. Our optimization results shed light on the significant impact of different factors on citric acid production. For instance, the addition of 0.4 g/L magnesium sulfate led to a maximum yield of 75%, while 2 g/L potassium ferrocyanide, added at 24 h, achieved a yield of 78%. Remarkably, ammonium oxalate, at a concentration of 10 g/L, resulted in a notable 77% yield. Conversely, the addition of calcium chloride exhibited negligible effects on citric acid production, with the control group yielding more at 78%. Our study underscores the potential for optimizing factors to enhance citric acid production by A. niger in submerged fermentation. These findings highlight the pivotal role of magnesium sulfate, potassium ferrocyanide, and ammonium oxalate in augmenting citric acid yields while emphasizing the minimal impact of calcium chloride. Ultimately, these insights contribute to advancing our understanding of microbial citric acid biosynthesis, providing valuable implications for industrial applications and future research endeavors.

The lemon genome and DNA methylome unveil epigenetic regulation of citric acid biosynthesis during fruit development.

Citric acid gives lemons their unique flavor, which impacts their sensory traits and market value. However, the intricate process of citric acid accumulation during lemon fruit growth remains incompletely understood. Here, we achieved a chromosomal-level genome assembly for the 'Xiangshui' lemon variety, spanning 364.85Mb across nine chromosomes. This assembly revealed 27 945 genes and 51.37% repetitive sequences, tracing the divergence from citron 2.85 million years ago. DNA methylome analysis of lemon fruits across different developmental stages revealed significant variations in DNA methylation. We observed decreased CG and CHG methylation but increased CHH methylation. Notably, the expression of RdDM pathway-related genes increased with fruit development, suggesting a connection with elevated CHH methylation, which is potentially influenced by the canonical RdDM pathway. Furthermore, we observed that elevated CHH DNA methylation within promoters significantly influenced the expression of key genes, critically contributing to vital biological processes, such as citric acid accumulation. In particular, the pivotal gene phosphoenolpyruvate carboxykinase (ClPEPCK), which regulates the tricarboxylic acid cycle, was strikingly upregulated during fruit development, concomitant with increased CHH methylation in its promoter region. Other essential genes associated with citric acid accumulation, such as the MYB transcription factor (ClPH1/4/5) and ANTHOCYANIN 1 (ClAN1), were strongly correlated with DNA methylation levels. These results strongly indicate that DNA methylation crucially orchestrates the metabolic synthesis of citric acid. In conclusion, our study revealed dynamic changes in DNA methylation during lemon fruit development, underscoring the significant role of DNA methylation in controlling the citric acid metabolic pathway.

A Review on Citric Acid Production by Yarrowia lipolytica Yeast: Past and Present Challenges and Developments

The biosynthesis of citric acid (CA) and its derivatives is of great interest due to its wide range of applications in various manufacturing sectors. The fungus Aspergillus niger is mainly used for the commercial production of CA, using sucrose and molasses as the primary carbon sources. Since the 1960s, intensive research has been underway to introduce Yarrowia lipolytica yeast as an alternative to traditional fungal technology. This review discusses the practical uses of CA and its derivatives. Also, the challenges and developments that have led to efficient and green CA synthesis technologies using Y. lipolytica are outlined. The nutrient medium requirements and the use of various carbon sources, encompassing pure substrates and industry, agriculture, and food waste are considered. Additionally, the choice and improvement of strain producers, including efficient mutagenesis, genetic modification, and screening methods, are discussed.

Costly effective bioleaching of valuable metals from low grade ore using Aspergillus nidulans

This research investigated the feasibility of employing organic acids, like citric acid, produced by Aspergillus nidulans MT355567 in a bioleaching process to recover uranium (U) from a low-grade rock sample. The optimal conditions for fungal growth and maximum citric acid (CA) synthesis across three distinct media were determined. The maximum citric acid concentration was produced on medium made from wheat bran (83%) and tea waste (77%). An investigation was carried out to see how citric acid and, by consequence, uranium bioleaching affinity, were affected by varying carbon sources, nitrogen sources, pH, temperature, incubation period, ore particle size, and the solid–liquid ratio. At 25 °C and a pH of 5.0, media containing 100 g/L of sucrose as a carbon source and peptone as a nitrogen source made the highest yield of citric acid and U bioleaching. Higher U bioleaching was achieved with ore particles 0.075 mm at a ratio of 2 g/L after only 30 min of contact with the fungal filtrate. Iron interference has a negative impact on uranium extraction. Interestingly, none of the conditions applied to enhance CA synthesis and U-bioleaching caused iron (Fe) dissolution. Based on these findings, it appears that bioleaching using A. nidulans MT355567 metabolic products is a promising economic and ecofriendly technology for extracting uranium from low-grade ore that might be adopted on a pilot scale.Graphical abstractsummarizing the experimental workflow for bioleaching of uranium from low-grade ore using citric acid produced by Aspergillus nidulans. The process involved optimizing A. nidulans growth and citric acid biosynthesis, evaluating factors influencing bioleaching activity of the acid metabolite solutions, and finally applying the optimized conditions to bioleach uranium from the ore sample. The schematic illustrates the key steps and the optimal condition for Aspergillus nidulans growth medium preparation using agricultural wastes, downstream application of metabolite synthesized for uranium bioleaching

Investigating the Usefulness of Wild Dioscorea esculenta (Wild Yam) Flour as Substrate for Citric Acid Biosynthesis

Background and Objective: The cost of substrate represents a significant percentage of the total biosynthesis cost of citric acid. There is need to continue searching for better and cheap underutilize starchy raw material for citric acid biosynthesis. Therefore, the objective of this research was to biologically synthesis citric acid in submerged fermentation of Aspergillus niger using wild Dioscorea esculenta (wild yam) tubers.
 Methodology: Submerged fermentation was conducted and the effect of Dioscorea esculenta concentrations, combined pretreatment and membrane permeability induction on citric acid biosynthesis were carried out.
 Results: Citric acid concentration increased as the concentration of the wild yam increased with maximum citric acid production of 10.5 ± 0.9 g/ l after 96 hours of fermentation. The overnight combined pretreatment with 0.3 % NaOH before pretreatment at 121oC for 20 minutes as one time combined pretreatment yielded 15. 0 ±1.2 g/l citric acid. Calcium chloride membrane induction produced 24.5 ± 1.3 g/l citric acid concentration respectively after 96 hours of fermentation.
 Conclusion: The result showed that wild Dioscorea esculenta tubers can be used as a substrate for the biosynthesis of citric acid. The biosynthesis of citric acid from Dioscorea esculenta tubers was enhanced by increased in concentration, combined pretreatment and membrane permeability induction.

Engineering a Phosphoketolase Pathway to Supplement Cytosolic Acetyl-CoA in Aspergillus niger Enables a Significant Increase in Citric Acid Production

Citric acid is widely used in the food, chemical and pharmaceutical industries. Aspergillus niger is the workhorse used for citric acid production in industry. A canonical citrate biosynthesis that occurred in mitochondria was well established; however, some research suggested that the cytosolic citrate biosynthesis pathway may play a role in this chemical production. Here, the roles of cytosolic phosphoketolase (PK), acetate kinase (ACK) and acetyl-CoA synthetase (ACS) in citrate biosynthesis were investigated by gene deletion and complementation in A. niger. The results indicated that PK, ACK and ACS were important for cytosolic acetyl-CoA accumulation and had significant effects on citric acid biosynthesis. Subsequently, the functions of variant PKs and phosphotransacetylase (PTA) were evaluated, and their efficiencies were determined. Finally, an efficient PK-PTA pathway was reconstructed in A. niger S469 with Ca-PK from Clostridium acetobutylicum and Ts-PTA from Thermoanaerobacterium saccharolyticum. The resultant strain showed an increase of 96.4% and 88% in the citrate titer and yield, respectively, compared with the parent strain in the bioreactor fermentation. These findings indicate that the cytosolic citrate biosynthesis pathway is important for citric acid biosynthesis, and increasing the cytosolic acetyl-CoA level can significantly enhance citric acid production.

Optimization of Medium Constituents for the Production of Citric Acid from Waste Glycerol Using the Central Composite Rotatable Design of Experiments

Citric acid is currently produced by submerged fermentation of sucrose with the aid of Aspergillus niger mold. Its strains are characterized by a high yield of citric acid biosynthesis and no toxic by-products. Currently, new substrates are sought for production of citric acid by submerged fermentation. Waste materials such as glycerol or pomace could be used as carbon sources in the biosynthesis of citric acid. Due to the complexity of the metabolic state in fungus, there is an obvious need to optimize the important medium constituents to enhance the accumulation of desired product. Potential optimization approach is a statistical method, such as the central composite rotatable design (CCRD). The aim of this study was to increase the yield of citric acid biosynthesis by Aspergillus niger PD-66 in media with waste glycerol as the carbon source. A mathematical method was used to optimize the culture medium composition for the biosynthesis of citric acid. In order to maximize the efficiency of the biosynthesis of citric acid the central composite, rotatable design was used. Waste glycerol and ammonium nitrate were identified as significant variables which highly influenced the final concentration of citric acid (Y1), volumetric rate of citric acid biosynthesis (Y2), and yield of citric acid biosynthesis (Y3). These variables were subsequently optimized using a central composite rotatable design. Optimal values of input variables were determined using the method of the utility function. The highest utility value of 0.88 was obtained by the following optimal set of conditions: waste glycerol-114.14 g∙L-1and NH4NO3-2.85 g∙L-1.

Biomanufacturing of food-grade citric acid and comprehensive utilization of its production wastewater

Citric acid is an important and multi-functional organic acid, which is widely used in food, medicine, chemical industry and other fields. It is the largest edible organic acid in the world. Aspergillus niger is an important strain for biosynthesis of citric acid, which is mainly produced by simultaneous saccharification and fermentation with starch raw materials. The reuse of citric acid wastewater is the development direction of comprehensive utilization of citric acid wastewater. In the future, with the combination of novel technology strategies such as gene editing technology and synthetic biology technology, the establishment of fermentation strategies with high yield, high yield, high production intensity and clean production is the development direction to realize efficient green manufacturing in citric acid industry.

Effect of Nitrogen Concentration on the Biosynthesis of Citric Acid, Protein, and Lipids in the Yeast Yarrowia lipolytica.

Yarrowia lipolytica yeast is well known to be able to synthesize citric acid (CA) in large amounts. This study deals with CA biosynthesis, the production of biomass, as well as the accumulation and composition of proteins and lipids in Y. lipolytica VKM Y-2373 grown in media with glucose at different concentrations of ammonium sulfate (from 2 to 10 g/L). It was found that these concentrations of nitrogen source are limiting for the growth of Y. lipolytica and that nitrogen deficiency is the main cause of CA excretion. At the high concentration of (NH4)2SO4 (10 g/L), the accumulation of cell biomass, biomass yield (YX/S), and protein concentration was higher than in the medium with 2 g/L ammonium sulfate by 4.3 times, 143%, and 5.1 times, respectively. CA was accumulated in meaningful quantities only in media containing 3–10 g/L (NH4)2SO4 with the maximum concentration of CA (99.9 g/L) at 4 g/L ammonium sulfate. Also of interest is the technological mode with 6 g/L (NH4)2SO4, which is characterized by high productivity (1.11 g/L × h). It should be noted that biomass contains large amounts of essential amino acids and unsaturated fatty acids and can be used in food biotechnologies and agriculture.

Identification of chitin synthase activator in Aspergillus niger and its application in citric acid fermentation.

The biosynthesis of citric acid (CA) using Aspergillus niger as a carrier is influenced by mycelium morphology, which is determined by the expression level of morphology-related genes. As a key component of the fungal cell wall, chitin content has an important effect on morphogenesis, and to investigate the effects of this on fermentation performance, we used RNA interference to knockdown chitin synthase C (CHSC) and chitin synthase activator (CHS3) to obtain the single-gene mutant strains A. niger chs3 and chsC and the double mutant A. niger chs3C. We found that the CA fermentation performance of the two single mutants was significantly better than that of the double mutant. The mutant A. niger chs3-4 exhibited CA production potential compared to that of the parent strain in scale-up fermentation; we determined certain characteristics of CA high-yielding strain fermentation pellets. In addition, when chsC alone was silenced, there was very little change in chs3 mRNA levels, whereas those of chsC were significantly reduced when only chs3 was silenced. As this may be because of a synergistic effect between chsC and chs3, and we speculated that the latent activation target of CHS3 is CHSC, our results confirmed this hypothesis. This study is the first application of a separation and combination silence strategy of chitin synthase and chitin synthase activator in the morphology of A. niger CA fermentation. Furthermore, it provides new insights into the method for the morphological study of A. niger fermentation and the interaction of homologous genes. KEY POINTS: • The function of chitin synthase C (chsC) and chitin synthase activator (chs3) is tightly interrelated. • Mycelial morphology was optimized by knockdown of CHS3, resulting in the overproduction of citric acid. • The separation and combination silence strategies are promising tools for the interaction of homologous housekeeping genes.

Understanding the influence of 2,4-dichlorophenoxyacetic acid and melatonin treatments on the sweet and acidic flavors and citric acid metabolism of ‘Olinda’ orange (Citrus sinensis (L.) Osbeck)

The regulatory mechanism by which 2,4-dichlorophenoxyacetic acid (2,4-D) and melatonin (MT) affect the sweet and acidic flavors of citrus fruit was investigated by comparing the fruit quality and gene expression related to citric acid metabolism of ‘Olinda’ oranges (Citrus sinensis (L.) Osbeck). 2,4-D increased the fruit firmness, total soluble solids, soluble sugar content and solid-acid ratio and decreased the respiration rate, titratable acidity and citric acid content during storage, and a similar but weaker effect was found in the MT treatment before 70 days of storage. 2,4-D and MT maintained postharvest quality and improved the fruit sweetness and acidity flavor of ‘Olinda’ Valencia oranges. Genes encoding of PEPC1/2/4 and CS1 in the citric acid biosynthesis pathway and Aco2/3, IDH2/3, ACLa2/β, PEPCK1 and FBPase2 in the citric acid degradation pathway were upregulated by 2,4-D or MT. However, genes related to citric acid degradation, such as GS2, Aco1, IDH1, GAD4/5 and SSADH in the γ-aminobutyric acid (GABA) pathway and ACLa1 and FBPase1 in the acetyl-CoA pathway, were oppositely regulated by 2,4-D and MT. Principal component analysis showed that the decrease in citric acid content was significantly negatively correlated with the expression of genes related to the GABA and acetyl-CoA degradation pathways. Moreover, the ACLα1/β and FBPase1 genes in the acetyl-CoA pathway might be the key genes causing differences in the acidity of ‘Olinda’ oranges among the three groups.

Influence of Gypsum-Containing Waste on Ammonia Binding in Animal Waste Composting

The possibility of using gypsum-containing waste–citrogypsum, which is a by-product of the chemical biosynthesis of citric acid, was considered as an additive for composting poultry manure from poultry farms that practice litter-free poultry keeping. The research was carried out on an experimental batch of 1500 tons. The production of the batch was carried out by mixing citrogypsum with a moisture content of 30% and litterless chicken manure with a moisture content of no more than 80% in a ratio of 1:2. The resulting mixture was placed on an open landfill in piles 3 m wide, 1 m high and 400 m long and was mixed twice with a compost turner. Further processing consisted of mixing the mass once every seven days. The controlled parameters were changes in humidity, temperature, pH and nitrogen content during composting. In the course of the experiment, it was found that the introduction of citrogypsum into the composition of the compost helps to optimize the moisture, temperature and pH of the mixture, and a decrease in ammonia emission to 87% was recorded, with an increase in nitrogen content of 2.4 times compared to the initial value. It was concluded that citrogypsum can be used in composting poultry waste to reduce volatilization of ammonia and preserve nutrients.

The Role of Hexokinase and Hexose Transporters in Preferential Use of Glucose over Fructose and Downstream Metabolic Pathways in the Yeast Yarrowia lipolytica

The development of efficient bioprocesses requires inexpensive and renewable substrates. Molasses, a by-product of the sugar industry, contains mostly sucrose, a disaccharide composed of glucose and fructose, both easily absorbed by microorganisms. Yarrowia lipolytica, a platform for the production of various chemicals, can be engineered for sucrose utilization by heterologous invertase expression, yet the problem of preferential use of glucose over fructose remains, as fructose consumption begins only after glucose depletion what significantly extends the bioprocesses. We investigated the role of hexose transporters and hexokinase (native and fructophilic) in this preference. Analysis of growth profiles and kinetics of monosaccharide utilization has proven that the glucose preference in Y. lipolytica depends primarily on the affinity of native hexokinase for glucose. Interestingly, combined overexpression of either hexokinase with hexose transporters significantly accelerated citric acid biosynthesis and enhanced pentose phosphate pathway leading to secretion of polyols (31.5 g/L vs. no polyols in the control strain). So far, polyol biosynthesis was efficient in glycerol-containing media. Moreover, overexpression of fructophilic hexokinase in combination with hexose transporters not only shortened this process to 48 h (84 h for the medium with glycerol) but also allowed to obtain 23% more polyols (40 g/L) compared to the glycerol medium (32.5 g/L).

Definitive Screening Design and Artificial Neural Network for Modeling a Rapid Biodegradation of Date Palm Fronds by a New Trichoderma sp. PWN6 into Citric Acid.

Generally, the bioconversion of lignocellulolytics into a new biomolecule is carried out through two or more steps. The current study used one-step bioprocessing of date palm fronds (DPF) into citric acid as a natural product, using a pioneer strain of Trichoderma harzianum (PWN6) that has been selected from six tested isolates based on the highest organic acid (OA) productivity (195.41 µmol/g), with the lowest amount of the released glucose. Trichoderma sp. PWN6 was morphologically and molecularly identified, and the GenBank accession number was MW78912.1. Both definitive screening design (DSD) and artificial neural network (ANN) were applied, for the first time, for modeling the bioconversion process of DPF. Although both models are capable of making accurate predictions, the ANN model outperforms the DSD model in terms of OA production, as ANN is characterized by a higher value of R2 (0.963) and validation R2 (0.967), and lower values of the RMSE (13.44), MDA (11.06), and SSE (9749.5). Citric acid was the only identified OA as was confirmed by GC-MS and UPLC, with a total of 1.5%. In conclusion, DPF together with T. harzianum PWN6 is considered an excellent new combination for citric acid biosynthesis, after modeling with artificial intelligence procedure.

Changes in the Content of Organic Acids and Expression Analysis of Citric Acid Accumulation-Related Genes during Fruit Development of Yellow (Passiflora edulis f. flavicarpa) and Purple (Passiflora edulis f. edulis) Passion Fruits.

Organic acids are key components that determine the taste and flavor of fruits and play a vital role in maintaining fruit quality and nutritive value. In this study, the fruits of two cultivars of passion fruit Yellow (Passiflora edulis f. flavicarpa) and purple (Passiflora edulis f. edulis) were harvested at five different developmental stages (i.e., fruitlet, green, veraison, near-mature and mature stage) from an orchard located in subtropical region of Fujian Province, China. The contents of six organic acids were quantified using ultra-performance liquid chromatography (UPLC), activities of citric acid related enzymes were determined, and expression levels of genes involved in citric acid metabolism were measured by quantitative real-time PCR (qRT-PCR). The results revealed that citric acid was the predominant organic acid in both cultivars during fruit development. The highest citric acid contents were observed in both cultivars at green stage, which were reduced with fruit maturity. Correlation analysis showed that citrate synthase (CS), cytosolic aconitase (Cyt-ACO) and cytosolic isocitrate dehydrogenase (Cyt-IDH) may be involved in regulating citric acid biosynthesis. Meanwhile, the PeCS2, PeACO4, PeACO5 and PeIDH1 genes may play an important role in regulating the accumulation of citric acid. This study provides new insights for future elucidation of key mechanisms regulating organic acid biosynthesis in passion fruit.

Biosynthesis of Hydroxy Citric Acid Mediated Zinc Nanoparticles and Its Antioxidant and Cytotoxic Activity

Aim: To evaluate biosynthesis of hydroxy citric acid mediated zinc nanoparticle and its antioxidant and cytotoxic activity
 Introduction: Hydroxy citric acid is a chemical that is found in fruit rinds of Garcinia cambogia, Garcinia indica, and Garcinia atroviridis. Hydroxy citric acid is used to improve exercise performance and weight loss. Hydroxy citric acid can cause nausea, digestive tract discomfort, and headache when used short-term. Long-term safety is unknown.Zinc oxide nanoparticles are nanoparticles of zinc oxide that have diameters less than 100 nanometers. Zinc nanoparticles have a large surface area relative to their size and high catalytic activity. The exact physical and chemical properties of zinc oxide nanoparticles depend on the different ways they are synthesized. zinc oxide nanoparticles have also been shown to exhibit strong protein adsorption properties, which can be used to modulate cytotoxicity, metabolism or other cellular responses.
 Materials and Methods: Evaluation of anti-oxidant was done by DPPH activity and evaluation of cytotoxic activity was done by brine shrimp lethality assay. Colour changes was noted in evaluation of cytotoxic activity
 Results: The antioxidant activity was exhibited well at higher doses when compared to standard, but there was only 40% of cytotoxicity achieved which indicates its potential.Cytotoxic activity reveals that the drug showed a very good result at a concentration of 25 ul it showed a 40% of lethality thereby showing that Hydroxycitric acid medicated zinc oxide nanoparticles shows a potent activity when compared to that with the standard.
 Conclusion: The study concludes that hydroxy citric acid shows good antioxidant and cytotoxic activity.

Transcriptome analysis of "Langmei" fruits and key enzyme genes structure and function prediction involved in citric acid biosynthesis

Prunus mume "Langmei" is a relict tree species, which fruit has good medicinal value. To understand the biosynthesis pathway of citric acid, the Illumina HiSeq XTen high-throughput sequencing technology was used to get the transcriptome from "Langmei". A total of 38 936 unigenes were obtained by assembling the fruit transcripts, of which 28 311 unigenes were successfully annotated in public databases, 15 193 unigenes were mapped to 265 KEGG metabolic pathways, and 18 908 unigenes were classified into 59 GO functional subclasses, 103 unigenes encoding 15 key enzymes involved in citric acid synthesis pathway were identified and analyzed. The structural model of citrate synthetase in "Langmei" showed that it was a homodimer and the secondary structure of each monomer was mainly composed of alpha helixes. Moreover, the residues in the active site of the citrate synthetase were highly conserved. This study provides a valuable resource for identifying candidate genes involved in the citric acid biosynthesis pathway, and will promote the development and sustainable utilization of genetic resources of "Langmei".

Overexpression of Citrate Synthase Increases Isocitric Acid Biosynthesis in the Yeast Yarrowia lipolytica

Yarrowia lipolytica is a non-conventional yeast producing valuable compounds, such as citric acids, from renewable raw materials. This study investigated the impact of citrate synthase overexpression on the biosynthesis of citric and isocitric acid in Y. lipolytica. Two transformants of Y. lipolytica A101.1.31 strain (efficient citric acid producer), overexpressing CIT1 or CIT2 gene (encoding proteins with citrate synthase activity), were constructed. The results revealed that overexpression of either of these genes enhances citrate synthase activity. Additionally, the cit1 knockout strain was unable to use propionate as the sole carbon source, which proves that CIT1 gene encodes a dual activity protein–citrate and 2-methylcitrate synthase. In the overexpressing mutants, a significant increase in isocitric acid biosynthesis was observed. Both CIT1 and CIT2 overexpressing strains produced citric and isocitric acid from vegetable oil in a ratio close to 1 (CA/ICA ratio for wild-type strain was 4.12).

Phytate hydrolysing activity of the Aspergillus niger L-4 micromycete strain

The aim of the study was to study the phytase synthesis capability of Aspergillus niger L-4 strain. The method for determining phytase activity is based on establishing the content of inorganic phosphates as a result of the action of phytase on the substrate under certain standard conditions by binding them with a vanadium-molybdenum reagent to form a coloured complex. The use of phytases for the hydrolysis of phytates in animal feed is important from the point of view of preserving the environment: when phytate complexes are destroyed, phosphorus is released, which performs an important structural and regulatory function, ensuring the normal development of bone and dental tissues and supporting their safety and integrity. Phosphoric acid is involved in the synthesis of kinases responsible for the normal course of chemical reactions in cells, in fat metabolism, as well as in the synthesis and breakdown of starch and glycogen. This reduces the release of undigested phosphorus into the environment. The object of the study consisted of native solutions obtained by culturing an industrial strain of acid-forming A. niger L-4 on various carbohydrate-containing media. The A. niger L-4 strain, previously selected at the All-Russian Scientific Research Institute of Food Additives for fermentation of molasses, has the ability to synthesise extracellular phytase. This paper presents the results of studies of phytase activity during the cultivation of A. niger L-4 on carbohydrate-containing media. It was found that in order components of the sucrose-mineral medium provide an elevated level of low-molecular-weight sugars necessary for increasing the productivity of phytase biosynthesis. Phytase activity in the native solution was shown to increase over 72 hours of fermentation to reach a value of 25.8±0.1 units/cm 3 . The phytase activity was 1.5 times higher than the fermentation process of a corn starch hydrolysate with a dextrose equivalent DE = 21±1 %, ensuring the productive biosynthesis of citric acid.

An Asymptotic Analysis of the Malonyl-CoA Route to 3-Hydroxypropionic Acid in Genetically Engineered Microbes

There has been recent interest in creating an efficient microbial production route for 3-hydroxypropionic acid, an important platform chemical. We develop and solve a mathematical model for the time-dependent metabolite concentrations in the malonyl-CoA pathway for 3-hydroxypropionic acid production in microbes, using a combination of numerical and asymptotic methods. This allows us to identify the most important targets for enzyme regulation therein under conditions of plentiful and sparse pyruvate, and to quantify their relative importance. In our model, we account for sinks of acetyl-CoA and malonyl-CoA to, for example, the citric acid cycle and fatty acid biosynthesis, respectively. Notably, in the plentiful pyruvate case we determine that there is a bifurcation in the asymptotic structure of the system, the crossing of which corresponds to a significant increase in 3-hydroxypropionic acid production. Moreover, we deduce that the most significant increases to 3-hydroxypropionic acid production can be obtained by up-regulating two specific enzymes in tandem, as the inherent nonlinearity of the system means that a solo up-regulation of either does not result in large increases in production. The types of issue arising here are prevalent in synthetic biology applications, and it is hoped that the system considered provides an instructive exemplar for broader applications.