Continous production of free gluconic acid by Gluconobacter suboxydans IFO 3290 immobilized by adsorption on ceramic honeycomb monolith: effect of reactor configuration on further oxidation of gluconic acid to keto-gluconic acid

Summary Gluconobacter suboxydans IFO 3290 was immobilized by adsorption on ceramic honeycomb monolith, and continuous production of free gluconic acid from 100 g/l glucose was carried out in one- and three-stage monolith reactors. Further oxidation of gluconic acid to keto-gluconic acid by the immobilized cells has been found to be more suppressed in the three-stage monolith reactor. This finding can be explained by the fact that, with the three-stage reactor, the opportunity to oxidize gluconic acid further was decreased because the residence time of the reaction mixture at glucose conversion above the threshold value was shorter.     

Continuous production of gluconic acid by immobilized Gluconobacter oxydans cell bioreactor

Summary Living Gluconobacter oxydans cells were attached on fibrous nylon carrier. Free gluconic acid was directly continuously produced in an aerated tubular immobilized-cell bioreactor for at least 6 months, with a volumetric productivity of at least 5 g/lh at 100 g/l substrate glucose and about 80 g/l product gluconic acid concentrations. The highest volumetric productivity in respect to glucose concentration was obtained with 175 g/l glucose, with about 120 g/l product gluconic acid level. With self-directing optimization procedure in respect to maximum product gluconic acid level, productivities as high as about 12–15 g/lh were obtained at relatively high substrate feed rate of 0.166 l/lh and relatively low aeration rate of 0.5 l/lmin. The highest glucose conversion of about 96% was obtained with a long residence time, at the lowest substrate feed rate used at a relatively low aeration rate, resulting however in a significant increase in ketogluconic acid production.     

Optimization of medium composition by response surface methodology for the production of tartaric acid by Gluconobacter suboxydans

The combined effect of sorbitol and yeast extract of the medium on tartaric acid production by Gluconobacter suboxydans, NCIM 2049, was studied in batch fermentation while keeping the temperature (30?°C) and pH (6.2) constant. Response surface methodology was used to obtain quadratic models for the production of tartaric acid. The multiple coefficients of regression between 0.8945 and 0.9820 was obtained during the process. The optimum medium composition comprising 20?kg/m³ sorbitol and 2?kg/m³ yeast extract was verified experimentally by observing the variation of cell mass and tartaric acid production with time.     

Optimization of temperature and initial pH and kinetic analysis of tartaric acid production by Gluconobacter suboxydans

Response surface methodology was applied for the simultaneous optimization of initial pH and temperature of fermentation for the production of tartaric acid by Gluconobacter suboxydans. A temperature of 32.8v°C and an initial pH of 6.05 was found to be most suitable. Kinetic analysis for the growth of Gluconobacter suboxydans and product formation was done and a logistic model was used to describe the growth of Gluconobacter suboxydans while the Leudeking-Piret model explained the product formation. Parameters of the model were evaluated and tartaric acid formation was found to be non-growth associated.     

Scale-down and optimization studies of the gluconic acid fermentation by Gluconobacter oxydans

To simulate production-scale conditions of gluconic acid fermentation by Gluconobacter oxydans, different experimental setups are presented in this study. From the determination of the time constants of a production-scale reactor, it can be concluded that mixing and oxygen transfer are the rate-limiting mechanisms. This results in oxygen concentration gradients which were simulated in a one-compartment reactor in which the oxygen concentration was fluctuated by a fluctuated gassing with air and nitrogen. It could be concluded that only very long periods of absence of oxygen (ca. 180 s) results in lower specific oxygen uptake rates by Gluconobacter oxydans. From scale-down studies carried out in a two-compartment system to simulate a production-scale reactor more accurately, it could be concluded that not only the residence time in the aerated part of the system is important, but the liquid flow in between the different parts of the reactor is also an essential parameter. It could also be concluded that the microorganisms are not influenced negatively by the fluctuated oxygen concentrations with respect to their maximal oxidation capacity. The two-compartment system can also be used for optimization experiments in which the "aerated" compartment was gassed with pure oxygen. From these experiments it was concluded that also a short residence of the cells at high oxygen concentrations diminished the growth and product formation rates. These experiments show the necessity of the scale-down experiments if optimization is carried out. The two-compartment system presented in this study is a very attractive tool for reliable scale-down experiments.     

L-Sorbose production by cells of the strain Gluconobacter suboxydans entrapped in a polyacrylamide gel

Summary It is shown, that bacteria of the strain Gluconobacter suboxydans entrapped in a polyacrylamide gel are capable to convert D-sorbitol to L-sorbose with a sufficiently high reaction rate. The kinetics of the studied process remains the same as it has been found for the case of free cells. Both semicontinuous and continuous patterns are accomplished.     

Production of free gluconic acid by cells ofGluconobacter oxydans

Summary Microbial oxidation of glucose to free gluconic acid by growingG.oxydans batch cultures was investigated. Kinetic data for the discussed process were obtained and attention was paid mainly to the influence of the initial glucose concentration and of the conversion degree on the course of the process. It was determined that relatively high maximum specific growth rates of about 0,39 h⁻¹ and gluconic acid volumetric productivities up to 53 mmol/h could be reached usingG.oxydans NBIMCC 1043 in runs without pH control. A maximum conversion degree of 90,4% was achieved.     

Synthesis of 2-keto-L-gulonic acid from gluconic acid by co-immobilized Gluconobacter oxydans and Corynebacterium sp.

2-Keto-L-gulonic acid was produced from gluconic acid using co-immobilized cells of Gluconobacter oxydans and Corynebacterium sp. with 2,5-diketo-D-gluconic acid. Gluconobacter oxydans and Corynebacterium sp. were entrapped together with polyvinylalcohol and alginate. 50 g/l glucose, 50 g/l gluconic acid, and the mixture of equal volume of 50 g/l glucose and 50 g/l gluconic acid were used as substrates. When the ratio of two cells was 1 to 1 with 100 mg cells/ml, the conversion of 2-KLG from gluconic acid was 38% (g/g). © Rapid Science Ltd. 1998     

代谢工程改造Gluconobacter suboxydans生产2-酮基-D-葡萄糖酸

2-酮基-D-葡萄糖酸是重要的抗氧化剂和食品添加剂——D-异抗坏血酸的重要前体。弱氧化葡糖酸杆菌(Gluconobacter suboxydans)具有丰富的周质空间氧化还原酶类,可将葡萄糖氧化为葡萄糖酸再氧化为2-酮基-D-葡萄糖酸。以提高2-酮基-D-葡萄糖酸的产量和减少副产物为目标,采用同源重组染色体修饰策略,将编码甘油脱氢酶的基因gldh置换为编码葡萄糖脱氢酶的基因gdh,将编码山梨醇脱氢酶的基因sdh置换为编码2-酮-D-葡萄糖酸脱氢酶的基因ga-2-dh。经PCR、酶活性显色及发酵产物HPLC检测验证表明:构建的工程菌株gdh和ga-2-dh基因被强化而gldh和sdh被敲除;使用10%的葡萄糖复合培养基,摇瓶发酵72h,工程菌2KGA3发酵液中没有副产物5-酮基-葡萄糖酸,2-酮基-D-葡萄糖酸的含量终浓度达到72.3 g/L,比野生菌株提高42.2g/L,工程菌和野生菌的2-D-KGA质量转化率分别为72.3%和30.1%,工程菌比野生菌提高1.4倍。构建获得的工程菌,不需要外加抗生素,可以保持稳定遗传,对于工业化规模生产具有一定优势,为获得可产业化显示的优势遗传资源打下了基础。     

Characterization of an insertion sequence, IS12528, from Gluconobacter suboxydans.

A novel insertion sequence element, IS12528, was found to be associated with inactivation of the alcohol dehydrogenase by insertion in the adhA gene, which encodes the primary dehydrogenase subunit of the three-component membrane-bound alcohol dehydrogenase complex in Gluconobacter suboxydans. Cloning and sequencing analyses revealed that IS12528 was 905 bp in length and had a terminal inverted repeat of 18 bp. In addition, IS12528 was found to generate a 3-bp duplication (TMA, where M represents C or A) at the inserted site upon transposition. IS12528 encoded one long product of 274 amino acids that was rich in basic amino acids. This protein showed significant homology with putative transposases of the IS1031 family isolated from Acetobacter xylinum, which belongs to another genus of acetic acid bacteria. IS12528-like sequences were distributed in a wide variety of acetic acid bacteria, as determined by Southern hybridization and PCR. These observations suggest that IS12528 is one of the insertion sequences that are responsible for genetic instability leading to deficiencies in various physiological properties in a variety of acetic acid bacteria.     

Microbiological production of gluconic acid by Egyptian mould fungi

Summary A local strain of Asp. phoenicis proved to be a good gluconic acid producer. Previous experiments carried out on this organism were extended to investigate the influence of some culture conditions on the acid production when the submerged technique was applied. These included the investigation of the influence of the nature and concentration of the nutritive constituents of the fermentation medium.     

Selective, high conversion of D-glucose to 5-keto-D-gluoconate by Gluconobacter suboxydans

Selective, high-yield production of 5-keto-D-gluconate (5KGA) from D-glucose by Gluconobacter was achieved without genetic modification. 5KGA production by Gluconobacter suffers byproduct formation of 2-keto-D-gluconate (2KGA). By controlling the medium pH strictly in a range of pH 3.5-4.0, 5KGA was accumulated with 87% conversion yield from D-glucose. The pH dependency of 5KGA formation appeared to be related to that of gluconate oxidizing activity.     

Selective,High Conversion of D-Glucose to 5-Keto-D-gluoconate by Gluconobacter suboxydans

Selective, high-yield production of 5-keto-D-gluconate (5KGA) from D-glucose by Gluconobacter was achieved without genetic modification. 5KGA production by Gluconobacter suffers byproduct formation of 2-keto-D-gluconate (2KGA). By controlling the medium pH strictly in a range of pH 3.5–4.0, 5KGA was accumulated with 87% conversion yield from D-glucose. The pH dependency of 5KGA formation appeared to be related to that of gluconate oxidizing activity.     

Biotechnological production of gluconic acid: future implications

Gluconic acid (GA) is a multifunctional carbonic acid regarded as a bulk chemical in the food, feed, beverage, textile, pharmaceutical, and construction industries. The favored production process is submerged fermentation by Aspergillus niger utilizing glucose as a major carbohydrate source, which accompanied product yield of 98%. However, use of GA and its derivatives is currently restricted because of high prices: about US$ 1.20–8.50/kg. Advancements in biotechnology such as screening of microorganisms, immobilization techniques, and modifications in fermentation process for continuous fermentation, including genetic engineering programmes, could lead to cost-effective production of GA. Among alternative carbohydrate sources, sugarcane molasses, grape must show highest GA yield of 95.8%, and banana must may assist reducing the overall cost of GA production. These methodologies would open new markets and increase applications of GA. Authors’ contributions OVS and RK are the sole contributors of this original review article. This review is based upon the published research in the area of gluconic acid fermentation.