Hanjie Ying

Name:

Organization: Nanjing University of Technology

Department: State Key Laboratory of Materials-Oriented Chemical Engineering, College of Life Science and Pharmaceutical Engineering

Title:

Chemicals
References

Determination of Metastable Zone Widths and the Primary Nucleation and Growth Mechanisms for the Crystallization of Disodium Guanosine 5′-Monophosphate from a Water–Ethanol System

Co-reporter:Fengxia Zou, Wei Zhuang, Jinglan Wu, Jingwei Zhou, PengPeng Yang, Qiyan Liu, Yong Chen, and Hanjie Ying

Industrial & Engineering Chemistry Research 2015 Volume 54(Issue 1) pp:137-145

Publication Date(Web):December 2, 2014

DOI:10.1021/ie504035j

Metastable zone widths (MSZWs) and induction times (tind’s) were measured by turbidity techniques during the antisolvent crystallization of disodium guanosine 5′-monophosphate (5′-GMPNa2). Measured MSZWs can be affected by numerous process parameters, including temperature, the agitation rate, and the antisolvent addition rate. An exponential equation was used to correlate the supersolubility and solubility data for different conditions, and to afford predictions of the MSZW values. Values of tind at different temperatures and mole fractions of water were assessed to determine the primary nucleation and growth mechanisms of 5′-GMPNa2 crystals in the water–ethanol system. The measured tind’s were then correlated using mononuclear and polynuclear mechanistic models. The fitting results identified the primary nucleation mechanism for 5′-GMPNa2 as polynuclear, which relates tind and the supersaturation for various growth mechanisms. The growth mechanism of 5′-GMPNa2 was found to be diffusion-controlled at all experimental temperatures.

Redirecting metabolic flux in Saccharomyces cerevisiae through regulation of cofactors in UMP production

Co-reporter:Yong Chen;Qingguo Liu;Xiaochun Chen

Journal of Industrial Microbiology & Biotechnology 2015 Volume 42( Issue 4) pp:577-583

Publication Date(Web):2015 April

DOI:10.1007/s10295-014-1536-y

Although it is generally known that cofactors play a major role in the production of different fermentation products, their role has not been thoroughly and systematically studied. To understand the impact of cofactors on physiological functions, a systematic approach was applied, which involved redox state analysis, energy charge analysis, and metabolite analysis. Using uridine 5′-monophosphate metabolism in Saccharomyces cerevisiae as a model, we demonstrated that regulation of intracellular the ratio of NADPH to NADP+ not only redistributed the carbon flux between the glycolytic and pentose phosphate pathways, but also regulated the redox state of NAD(H), resulting in a significant change of ATP, and a significantly altered spectrum of metabolic products.

A mild and highly efficient laccase-mediator system for aerobic oxidation of alcohols

Co-reporter:Chenjie Zhu, Zhi Zhang, Weiwei Ding, Jingjing Xie, Yong Chen, Jinglan Wu, Xiaochun Chen and Hanjie Ying

Green Chemistry 2014 vol. 16(Issue 3) pp:1131-1138

Publication Date(Web):03 Dec 2013

DOI:10.1039/C3GC42124D

With the aid of the highly active nitroxyl radical AZADO (2-azaadamantane N-oxyl), a simple method for the aerobic catalytic oxidation of alcohols is presented. The oxidations could typically proceed under practical ambient conditions (room temperature, air atmosphere, no moisture effect, metal-free, etc.) with a broad generality of the alcohol substrates, and especially for the oxidation of complex and highly functionalized alcohols. An ionic mechanism is proposed for the present system.

Recovery of Acetoin from the Ethanol–Acetoin–Acetic Acid Ternary Mixture Based on Adsorption Methodology Using a Hyper-Cross-Linked Resin

Co-reporter:Jinglan Wu, Xu Ke, Lili Wang, Renjie Li, Xudong Zhang, Pengfei Jiao, Wei Zhuang, Yong Chen, and Hanjie Ying

Industrial & Engineering Chemistry Research 2014 Volume 53(Issue 31) pp:12411-12419

Publication Date(Web):July 14, 2014

DOI:10.1021/ie502105q

The separation performances, in terms of adsorption selectivity, desorption, and regeneration of an innovative hyper-cross-linked resin (HD-02) for recovery of acetoin from the ethanol–acetoin–acetic acid ternary mixture were explored in this work. The competitive adsorption behaviors of the ternary mixture were determined experimentally. The results showed the HD-02 resin had a good adsorption selectivity toward acetoin over acetic acid and ethanol. Subsequently, the desorption behaviors in terms of desorption isotherms and kinetics were systematically investigated. Using ethanol as a desorbent, the recovery could be achieved as high as 98% in the batch desorption experiments. The Fick model was adopted to simulate the desorption process. The simulation results revealed that the intraparticle diffusion was the rate-limiting step and the obtained effective diffusivities (1.530 × 10–9 m2/min) were independent of the acetoin concentrations. In the end of this work, three cycles of adsorption–desorption–regeneration operations confirmed good reproducibility of the resin for the attainment of acetoin.

Control of glycolytic flux in directed biosynthesis of uridine-phosphoryl compounds through the manipulation of ATP availability

Co-reporter:Yong Chen;Qingguo Liu;Xiaochun Chen;Jinglan Wu

Applied Microbiology and Biotechnology 2014 Volume 98( Issue 15) pp:6621-6632

Publication Date(Web):2014 August

DOI:10.1007/s00253-014-5701-z

Adenosine triphosphate (ATP), the most important energy source for metabolic reactions and pathways, plays a vital role in control of metabolic flux. Considering the importance of ATP in regulation of the glycolytic pathway, the use of ATP-oriented manipulation is a rational and efficient route to regulate metabolic flux. In this paper, a series of efficient ATP-oriented regulation methods, such as changing ambient temperature and altering reduced nicotinamide adenine dinucleotide (NADH), was developed. To satisfy the different demand for ATP at different phases in directed biosynthesis of uridine-phosphoryl compounds, a multiphase ATP supply regulation strategy was also used to enhance to yield of target metabolites.

Experimental measurement and modelling of solubility of inosine-5′-monophosphate disodium in pure and mixed solvents

Co-reporter:Fengxia Zou, Wei Zhuang, Jinglan Wu, Jingwei Zhou, Qiyan Liu, Yong Chen, Jingjing Xie, Chenjie Zhu, Ting Guo, Hanjie Ying

The Journal of Chemical Thermodynamics 2014 Volume 77() pp:14-22

Publication Date(Web):October 2014

DOI:10.1016/j.jct.2014.04.023

•Solubility of 5′-IMPNa2 in various solvents was studied for the first time.•The solubility could be ranked as follows: water > methanol > ethanol > acetone.•Modified Apelblat equation gave the best correlating results.•Mixing Gibbs free energies, enthalpies, and entropies were predicted.•Solubility data and equations can optimise the crystallization conditions.The solubility of biological chemicals in solvents provide important fundamental data and is generally considered as an essential factor in the design of crystallization processes. The equilibrium solubility data of inosine-5′-monophosphate disodium (5′-IMPNa2) in water, methanol, ethanol, acetone, as well as in the solvent mixtures (methanol + water, ethanol + water, acetone + water), were measured by an isothermal method at temperatures ranging from (293.15 to 313.15) K. The measured data in pure and mixed solvents were then modelled using the modified Apelblat equation, van’t Hoff equation, λh equation, ideal model and the Wilson model. The modified Apelblat equation showed the best modelling results, and it was therefore used to predict the mixing Gibbs free energies, enthalpies, and entropies of 5′-IMPNa2in pure and binary solvents. The positive values of the calculated partial molar Gibbs free energies indicated the variations in the solubility trends of 5′-IMPNa2. Water and ethanol (in the binary mixture with water) were found to be the most effective solvent and anti-solvent, respectively.Graphical abstract

Solubility of disodium cytidine 5′-monophosphate in different binary mixtures from 288.15 K to 313.15 K

Co-reporter:Jin Yu, Tianle Ma, An Li, Xiaochun Chen, Yong Chen, Jingjing Xie, Jinglan Wu, Hanjie Ying

Thermochimica Acta 2013 Volume 565() pp:1-7

Publication Date(Web):10 August 2013

DOI:10.1016/j.tca.2013.04.018

•Solubility of 5′-CMPNa2 in different systems was measured the first time.•Experimental data were correlated by CNIBS/Redlich–Kister model and Apelblat model.•Good agreement has been observed between the calculated and the experimental data.•Enthalpy and entropy were calculated by the van’t Hoff equation and Gibbs equation.The solubility of disodium cytidine 5′-monophosphate (5′-CMPNa2) in methanol + water and ethanol + water binary mixtures was measured experimentally at the temperatures ranging from 288.15 to 313.15 K. The results showed that the solubility of 5′-CMPNa2 increased with the increasing of temperature and the mole fraction of water in different binary mixtures. The (CNIBS)/Redlich–Kister model and the semi-empirical Apelblat model were applied for the prediction of the experimental data. Both models could give satisfactory simulation results. In addition, the thermodynamic properties of the dissolution process such as Gibbs energy, enthalpy, and entropy were calculated using the van’t Hoff equation and the Gibbs equation. The results indicated that the dissolution process was endothermic.

Enhancement of adenosine production by Bacillus subtilis CGMCC 4484 through metabolic flux analysis and simplified feeding strategies

Co-reporter:Xiaochun Chen;Chunwei Zhang;Jian Cheng

Bioprocess and Biosystems Engineering 2013 Volume 36( Issue 12) pp:1851-1859

Publication Date(Web):2013 December

DOI:10.1007/s00449-013-0959-6

The objective of this research was to understand how the initial glucose concentration influences adenosine (AR) production and metabolic flux shift on the cultivation of Bacillus subtilis CGMCC 4484. Experiments confirmed that initial glucose concentration affects cell growth, AR production and metabolites, significantly. The flux distribution at the key nodes of glucose-6-phosphate (G6P), pyruvate (PYR) and acetyl coenzyme-A (AcCoA) could be affected by changing the glucose concentration. Based on kinetic analysis of specific rates, the low-glucose concentration was better for both cell growth and AR production during the first 12 h. However, the high-glucose concentration was more favorable for AR formation after 18 h. Furthermore, different simplified feeding strategies were designed to achieve higher AR accumulation. The final AR concentration of 15.60 g L−1 was achieved when an optimized constant-feeding strategy was used, which was 21.02 % higher than batch fermentation. This was the first time to investigate the regulation of the glucose metabolism of AR-producing B. subtilis.

Production of butanol from glucose and xylose with immobilized cells of Clostridium acetobutylicum

Co-reporter:Yong Chen;Tao Zhou;Dong Liu;An Li;Songbo Xu

Biotechnology and Bioprocess Engineering 2013 Volume 18( Issue 2) pp:234-241

Publication Date(Web):2013 April

DOI:10.1007/s12257-012-0573-5

Pretreated cotton towels were used as carriers to immobilize Clostridium acetobutylicum CGMCC 5234 cells for butanol or ABE production from glucose and xylose. Results showed that cell immobilization was a promising method to increase butanol concentration, yield and productivity regardless of the sugar sources compared with cell suspension. In this study, a high butanol concentration of 10.02 g/L with a yield of 0.20 g/g was obtained from 60 g/L xylose with 9.9 g/L residual xylose using immobilized cells compared with 8.48 g/L butanol and a yield of 0.141 g/g with 20.2 g/L residual xylose from 60 g/L xylose using suspended cells. In mixed-sugar fermentation (30 g/L glucose plus 30 g/L xylose), the immobilized cultures produced 11.1 g/L butanol with a yield of 0.190 g/g, which were 28.3% higher than with suspended cells (8.65 g/L) during which 30 g/L glucose was utilized completely using both immobilized and suspended cells while 3.46 and 13.1 g/L xylose maintained untilized for immobilized and suspended cells, respectively. Based on the results, we speculated that immobilized cells showed enhanced tolerance to butanol toxicity and the cultures preferred glucose to xylose during ABE fermentation. Moreover, the cultures showed obvious difference when grown between high initial concentrations of glucose and those of xylose. Repeated-batch fermentations from glucose with immobilized cells showed better long-term stability than from xylose. At last, the morphologies of free and immobilized cells adsorbed on pretreated cotton towels during the growth cycle were examined by SEM.

Experimental and modeling studies on the sorption breakthrough behaviors of butanol from aqueous solution in a fixed-bed of KA-I resin

Co-reporter:Xiaoqing Lin;Renjie Li;Qingshi Wen

Biotechnology and Bioprocess Engineering 2013 Volume 18( Issue 2) pp:223-233

Publication Date(Web):2013 April

DOI:10.1007/s12257-012-0549-5

Removal of biobutanol from acetone-butanolethanol (ABE) fermentation broth can be achieved by fixed-bed sorption by means of KA-I resin, and the relevant breakthrough curves would provide much valuable information to help design a continuous fixed-bed sorption process in field application. In the present study, the effects of several important design parameters, i.e., initial butanol concentration (Cf: 3.0 ∼ 30.0 g/L), inlet flow rate (Qf: 0.5 ∼ 5.5 mL/min) and adsorbent bed height (Z: 4.2 ∼ 18.0 cm), on the adsorption breakthrough curves of KA-I resin in a fixed-bed column were investigated. It was found that the amount of adsorbed butanol at breakthrough point was increased with an increase in the value of Cf and Z; and with decrease in the value of Qf. However, the maximum sorption capacities of butanol at saturated point were basically unchanged. Three well-established fixed-bed adsorption models, namely Thomas, Yoon-Nelson and Adams-Bohart, were applied to predict the breakthrough curves and to determine the characteristic parameters of fixed-bed column, which are the basis for the process design at a real scale. Good agreement between the theoretical breakthrough curves and the experimental result were observed using Thomas and Yoon-Nelson models.

Experimental Determination of Metastable Zone Width, Induction Period, and Primary Nucleation Kinetics of Cytidine 5′-Monophosphate Disodium Salt in an Ethanol–Aqueous Mixture

Co-reporter:Jin Yu, An Li, Xiaochun Chen, Yong Chen, Jingjing Xie, Jinglan Wu, and Hanjie Ying

Journal of Chemical & Engineering Data 2013 Volume 58(Issue 5) pp:1244-1248

Publication Date(Web):April 9, 2013

DOI:10.1021/je4000263

A systematic investigation was carried out to determine the metastable zone width (MSZW) and induction period values of cytidine 5′-monophosphate disodium salt (5′-CMPNa2) in an ethanol + water mixture solution. Experimental determination of the MSZW was performed using a laser method. The two factors solution temperature and agitation were evaluated in this study to measure the MSZW. It was observed that the MSZWs decreased when the agitation levels increased and were little affected by solution temperature. The induction period was combined with the classical theory of nucleation. The various critical nucleation parameters, such as the interfacial tension (γ), the radius of critical nucleus (r*), the critical free energy of nucleus (ΔG*) and the molecular number of critical nucleus (i*) were evaluated in the controlled nucleation condition. The critical nucleation parameters varied with increased temperature, and the nucleation rate increased with increased supersaturation.

Production of cyclic adenosine-3′,5′-monophosphate by whole cell catalysis using recombinant Escherichia coli overexpressing adenylate cyclase

Co-reporter:Nan Li;Ying He;Yong Chen;Xiaochun Chen

Korean Journal of Chemical Engineering 2013 Volume 30( Issue 4) pp:913-917

Publication Date(Web):2013 April

DOI:10.1007/s11814-012-0202-1

Adenylate cyclase (EC 4.6.1.1) catalyzes the formation of cyclic adenosine-3′,5′-monophosphate (cAMP) from adenosine 5′-triphosphate (ATP). Recombinant Escherichia coli overexpressing adenylate cyclase was used to synthesize cAMP by whole cell catalysis. Some key parameters were examined during the catalytic process, while pH and Mg2+ were found to influence cAMP production significantly. Optimum conditions were pH 8.52 and 30 °C with 77.2 mM Mg2+ in 100 mM Tris-HCl buffer, including 0.25% Triton-X 100 as detergent and 30 mM pyruvate sodium as enzyme activator for 6 h. 14.93 g/L of cAMP was produced with a conversion rate of 91.5%. The current work provided a potential way for the industrial production of cAMP.

Adsorption of butanol from aqueous solution onto a new type of macroporous adsorption resin: Studies of adsorption isotherms and kinetics simulation

Co-reporter:Xiaoqing Lin;Jinglan Wu;Jiansheng Fan;Wenbin Qian;Xiqun Zhou;Chen Qian;Xiaohong Jin;Lili Wang;Jianxin Bai

Journal of Chemical Technology and Biotechnology 2012 Volume 87( Issue 7) pp:924-931

Publication Date(Web):

DOI:10.1002/jctb.3701

Abstract

BACKGROUND: Owing to the rapid depletion of petroleum fuel, the production of bio-butanol has attracted much attention. However, low butanol productivity severely limits its potential industrial application. It is important to establish an approach for recovering low-concentration butanol from fermentation broth. Experiments were conducted using batch adsorption mode under different conditions of initial butanol concentration and temperature. Batch adsorption data were fitted to Langmuir and Freundlich isotherms and the macropore diffusion, pseudo-first- and second-order models for kinetic study.

RESULTS: The maximum adsorption capacity of butanol onto KA-I resin increase with increasing temperature, ranged from 139.836 to 304.397 mg g⁻¹. The equilibrium adsorption data were well fitted by the Langmuir isotherm. The adsorption kinetics was more accurately represented by the macropore diffusion model, which also clearly predicted the intraparticle distribution of the concentration. The effective pore diffusivity (D_p) was dependent upon temperature, but independent of initial butanol concentration, and was 0.251 × 10⁻¹⁰, 0.73 × 10⁻¹⁰, 1.32 × 10⁻¹⁰ and 4.31 × 10⁻¹⁰ m² s⁻¹ at 283.13, 293.13, 303.13 and 310.13 K, respectively.

CONCLUSION: This work demonstrates that KA-I resin is an efficient adsorbent for the removal of butanol from aqueous solutions and available for practical applications for future in situ product recovery of butanol from ABE fermentation broth. Copyright © 2012 Society of Chemical Industry

Bi-stage control of dissolved oxygen to enhance cyclic adenosine monophosphate production by Arthrobacter A302

Co-reporter:Lei Li;Xiaochun Chen;Jian Cheng;Chunwei Zhang

Bioprocess and Biosystems Engineering 2012 Volume 35( Issue 8) pp:1281-1286

Publication Date(Web):2012 October

DOI:10.1007/s00449-012-0715-3

Experiments confirmed dissolved oxygen (DO) definitely affects cyclic adenosine monophosphate (cAMP) production by Arthrobacter A302. Production of cAMP by batch fermentation was investigated under various DO conditions. A two-stage DO control strategy was proposed to achieve optimal production of cAMP based on the kinetic analysis: the DO level was controlled at 40% during the first 18 h and then switched to 30%. Relatively high cAMP production (9.9 g L−1) was achieved by applying this strategy. The cAMP productivity (0.14 g L−1 h−1) was also successfully improved by 85.1, 59.3, 15.1 and 28.0%, compared to cases in which DO was uncontrolled or DO levels were held at 20, 30 and 40%, respectively. This is the first report of the use of a two-stage DO control strategy in cAMP production, and it was verified to be an effective method for enhancing the cAMP yield via this strain.

Adaptation of Glycolysis and Growth to Acetate in Sporolactobacillus sp. Y2-8

Co-reporter:Dong Liu;Yong Chen;An Li;Ting Zhao;Tao Zhou

Applied Biochemistry and Biotechnology 2012 Volume 168( Issue 2) pp:455-463

Publication Date(Web):2012 September

DOI:10.1007/s12010-012-9789-2

Exogenous addition of a low concentration of acetate (2 g/L sodium acetate) effectively decreased acetic acid excretion and lowered the ATP content in Sporolactobacillus sp. Y2-8 without any growth defect although the acetate could not be utilized at an initial glucose concentration of 150 g/L. This induced an enhanced glycolytic flux with increased specific activities of hexokinase and phosphofructokinase, probably to compensate for the lowered efficiency of ATP production. However, with increasing concentrations (5 g/L sodium acetate), acetate was utilized first before being produced again, causing a growth lag at the transition. Glucose consumption was also reduced at high acetate concentrations, resulting in decreased d-lactic acid production. These results demonstrate that acetate plays a significant role in regulating glycolysis and growth of Sporolactobacillus.

Enhanced uridine 5′-monophosphate production by whole cell of Saccharomyces cerevisiae through rational redistribution of metabolic flux

Co-reporter:Dong Liu;Yong Chen;An Li;Jingjing Xie

Bioprocess and Biosystems Engineering 2012 Volume 35( Issue 5) pp:729-737

Publication Date(Web):2012 June

DOI:10.1007/s00449-011-0653-5

A whole-cell biocatalytic process for uridine 5′-monophosphate (UMP) production from orotic acid by Saccharomyces cerevisiae was developed. To rationally redistribute the metabolic flux between glycolysis and pentose phosphate pathway, statistical methods were employed first to find out the critical factors in the process. NaH2PO4, MgCl2 and pH were found to be the important factors affecting UMP production significantly. The levels of these three factors required for the maximum production of UMP were determined: NaH2PO4 22.1 g/L; MgCl2 2.55 g/L; pH 8.15. An enhancement of UMP production from 6.12 to 8.13 g/L was achieved. A significant redistribution of metabolic fluxes was observed and the underlying mechanism was discussed.

Kinetic models of ribonucleic acid fermentation and continuous culture by Candida tropicalis no.121

Co-reporter:Bingbing Li;Xiaochun Chen;Huajing Ren;Lei Li

Bioprocess and Biosystems Engineering 2012 Volume 35( Issue 3) pp:415-422

Publication Date(Web):2012 March

DOI:10.1007/s00449-011-0580-5

During ribonucleic acid fermentation, the fermentative processes were researched at pH controlled at 4.0 and under natural conditions. Unstructured models in a 50-L airlift fermentor were established for batch RNA production at pH 4.0 using the Verhulst equation for microbial growth, the Luedeking–Piret equation for product formation and a Luedeking–Piret-like equation for substrate uptake. Parameters of the kinetic models were determined using origin 7.5. Based on the models estimated above, another batch fermentation experiment was conducted in a 300-L airlift fermentor, which demonstrated that the models could be useful for RNA production on an industrial scale. Additionally, continuous fermentation based on kinetic models was proposed to make full use of substrates and reduce the cost of waste water treatment. As a result, although the DCW and RNA concentration were 11.5 and 1.68 g L−1, which were lower than that of batch fermentation, the sugar utilization increased by 14.3%, while the waste water decreased by more than 90%.

Separation of d-lactic acid from aqueous solutions based on the adsorption technology

Co-reporter:Jinglan Wu, Yanan Hu, Jingwei Zhou, Wenbin Qian, Xiaoqing Lin, Yong Chen, Xiaochun Chen, Jingjing Xie, Jianxin Bai, Hanjie Ying

Colloids and Surfaces A: Physicochemical and Engineering Aspects 2012 Volume 407() pp:29-37

Publication Date(Web):5 August 2012

DOI:10.1016/j.colsurfa.2012.04.051

A novel polar macroporous adsorption resin (AX-1) has been applied for separation of d-lactic acid (DLA) from aqueous solutions in a fixed-bed chromatographic column. The adsorption equilibrium and kinetics were investigated experimentally. A column model was subsequently established and validated by comparison of the simulation results to the experimental data obtained at different operating conditions in terms of feed flow rates and initial concentrations as well as bed heights. At the end of this work, the recycling studies were carried out. It was found that the saturated resin could be washed out DLA completely and repeated use by 353 K hot water. The results show that the AX-1 resin has a high adsorption capacity to DLA while no retention to sulfate ion. Based on this kind of resin, a green-production refining process could be established since only water is added in the entire separation process.Graphical abstractHighlights► A novel polar macroporous resin was offered with good properties. ► A green refining process was proposed since only water was added. ► A model was established to design the d-lactic acid adsorption process.

Production of cyclic adenosine monophosphate by Arthrobacter sp. A302 using fed-batch fermentation with pH-shift control

Co-reporter:Jiaming Cao;Xiaochun Chen;Huajing Ren

World Journal of Microbiology and Biotechnology 2012 Volume 28( Issue 1) pp:121-127

Publication Date(Web):2012 January

DOI:10.1007/s11274-011-0799-y

The production of cyclic adenosine monophosphate (cAMP) by Arthrobacter sp. A302 was studied in a 5 L stirred tank fermentor under a range of pH values (6.5–8.0) and glucose feeding rates. In batch fermentation under a controlled pH, the optimum pH for cell growth was 7.5 with dry cell density (X) of 11.43 g L, and the optimum pH for cAMP accumulation was 7.0 with cAMP concentration of 7.41 g L. In order to achieve the high X and cAMP yield simultaneously, a pH-shift control strategy was proposed based on kinetic analysis of specific cell growth rate (μ) and specific cAMP formation rate (qs). In this method, pH was controlled to 7.0 for the first 30 h of fermentation, and then subsequently shifted to 7.5 and maintained until the end of the process. Application of this approach significantly enhanced the cAMP concentration. Thereafter, cAMP production was further improved by combining the above-mentioned pH-control system and fed-batch process with glucose at a constant feeding rate of 1.0 g L−1 h−1. Under optimum conditions, the final cAMP production was 10.87 g L, which is 110.0, 46.7, and 27.7% higher than that of the pH-uncontrolled, pH-controlled, and pH-shift controlled methods, respectively.

Cloning, expression, and characterization of an adenylate cyclase from Arthrobacter sp. CGMCC 3584

Co-reporter:Ying He;Nan Li;Yong Chen;Xiaochun Chen

Applied Microbiology and Biotechnology 2012 Volume 96( Issue 4) pp:963-970

Publication Date(Web):2012 November

DOI:10.1007/s00253-012-3890-x

The cya gene encoding adenylate cyclase was cloned from Arthrobacter sp. CGMCC 3584 by thermal asymmetric interlaced PCR for the first time. It exhibited an open reading frame containing 1,125 bp and encoding 374 amino acids. Amino acid sequence analysis showed that this enzyme was a class III adenylate cyclase. Expression of the cya gene was carried out in Escherichia coli Rosetta, and purification was performed via Ni2+-NTA agarose gel column. SDS-PAGE indicated that the molecular mass of the recombinant adenylate cyclase was 45 kDa. The Vmax and Km were determined to be 5.06 μmol/min/mg and 7.56 mM, respectively. The optimum pH and temperature were 8.0 and 35 °C. Several divalent metal ions were found to activate the enzyme to different extents, and the maximal specific activity reached 3.04 μmol/min/mg when 50 mM Mg2+ was added. This was the first report of the cloning of an adenylate cyclase gene from Arthrobacter sp.

Adsorption Thermodynamics and Kinetics of Uridine 5′-Monophosphate on a Gel-Type Anion Exchange Resin

Co-reporter:Xiqun Zhou, Jiansheng Fan, Nan Li, Wenbin Qian, Xiaoqing Lin, Jinglan Wu, Jian Xiong, Jianxin Bai, and Hanjie Ying

Industrial & Engineering Chemistry Research 2011 Volume 50(Issue 15) pp:9270-9279

Publication Date(Web):June 16, 2011

DOI:10.1021/ie101721a

The adsorption behavior of uridine 5′-monophosphate (UMP) on a gel-type anion-exchange resin SD3 at different temperatures was investigated by the batch method. The dissociation equilibrium of UMP in aqueous solution has been studied. Effects of solution pH, contact time, and initial concentrations of UMP on the adsorption have been discussed. Adsorption equilibrium data had been fitted to three different isotherms including Langmuir, Freundlich, and Sips isotherms, which have been widely used in biosorption processes, and the accuracy for all models has been evaluated by the residual-root-mean-square error. For adsorption kinetics, the adsorption rate of UMP on the resin was interpreted by the Fick model, first- and second-order kinetic models, and the adsorption process was found to be well represented by the Fick model. The solution diffusivities of UMP at different temperatures were estimated by the Wilke-Chang equation. The sorption process was found to be controlled by the intraparticle diffusion. The Fick model and Sips isotherm were chosen to simulate the concentration diffusion of UMP on SD3 resin during the adsorption process. An intraparticle two-dimensional profile of SD3 resin at 0, 1, 4, 15, and 25 min for 293.15, 303.15, and 313.15 K was shown with a satisfactory description of the adsorption process. The thermodynamic parameters such as Gibbs free energy and enthalpy and entropy changes were calculated, and the values indicated that the adsorption process of UMP on SD3 resin was spontaneous and endothermic.

Conformers and hydrogen bonds in cytidine 5′-diphosphocholine sodium single crystals grown from a mixture of ethanol and water

Co-reporter:Zhenxing Du, Yanan Hu, Pei Wang, Jingwei Zhou, Jian Xiong, Hanjie Ying, Jianxin Bai

Journal of Molecular Structure 2011 Volume 985(2–3) pp:227-232

Publication Date(Web):31 January 2011

DOI:10.1016/j.molstruc.2010.10.048

The molecular structure of cytidine 5′-diphosphocholine sodium (CDPC) grown from a mixture of ethanol and water was determined by X-ray diffraction (XRD). CDPC was found to have an orthorhombic structure with confirmed lattice parameters of a = 6.978 Å, b = 12.406 Å and c = 29.326 Å. This nucleotide coenzyme was highly folded and net-like. Each crystallographic unit consisted of one sodium atom, one pyrophosphate group, one cytosine group, one coordinated water molecule, one pentose molecule, and three lattice water molecules. The interspaces of neighboring CDPC molecules were filled with water molecules and methyl groups. Although the coordinated water was connected to sodium atoms, the lattice water molecules formed chair-shaped water hexamers. The hydrogen bonds which played an important role in maintaining the structure included OH···O, NH···O and CH···O and ranged in length from 2.682 (17) to 3.349 (17) Å. Fourier transform infrared spectroscopy (FTIR) showed a broad absorption in the 400–2000 cm−1 region characteristic of short hydrogen bonds. So for industrial crystallization, methods which could eliminate the influence of hydrogen bonds should be taken, and it would be beneficial for the process of crystallization.

Dynamic mathematical models of batch experiments and fed-batch cultures for cyclic adenosine monophosphate production by Arthrobacter A302

Co-reporter:Lei Li;Xiaochun Chen;Huajing Ren

World Journal of Microbiology and Biotechnology 2011 Volume 27( Issue 10) pp:2379-2385

Publication Date(Web):2011 October

DOI:10.1007/s11274-011-0707-5

A glucose utilizing strain, Arthrobacter A302 was used for cyclic adenosine monophosphate (cAMP) production in batch modes. The non-structured model in a 5 l stirred tank bioreactor for understanding, controlling, and optimizing the fermentation process was proposed using the logistic equation for microbial growth, the Luedeking-Piret equation for product formation and Luedeking-Piret-like equation for substrate uptake, respectively. The production of cAMP was a mixed-growth-associated pattern. Based on model prediction, a comparison of calculated value using the parameters evaluated above with another experimental data in 30 l bioreactor was used to test the model. The results predicted from the model were in good agreement with the experimental observations in 30 l bioreactor, which demonstrated that the model might be useful for the development and optimization of production of cAMP in industrial scale. Based on estimated kinetic parameters, three different fed-batch modes, constant rate and intermittent (once and repeated), were adopted in order to obtain more cAMP accumulation. Furthermore, the final production of cAMP reached 11.24 g l−1 after 72 h incubation using three stages feeding strategy. In particular, the cAMP productivity (0.156 g l−1 h−1) was successfully improved by 22.83, 11.43 and 9.86%, respectively, compared with the modes of the batch, constant rate fed-batch and intermittent fed-batch once.

Determination of optimal conditions for ribonucleic acid production by Candida tropicalis no. 121

Co-reporter:Huajing Ren;Xiaochun Chen;Jiaming Cao;Lei Li

Korean Journal of Chemical Engineering 2011 Volume 28( Issue 8) pp:

Publication Date(Web):2011 August

DOI:10.1007/s11814-011-0013-9

The experiments were based on multivariate statistical concepts, and response surface methodology (RSM) was applied to optimize the fermentation medium for the production of ribonucleic acid (RNA) by Candida tropicalis no. 121. The process involved the individual adjustment and optimization of various medium components at shake flask level. The two-level Plackett-Burman (PB) design was used to screen the medium components, which significantly influenced RNA production. Among seven variables, the concentrations of molasses, ZnSO4, and H3PO4 were found to be the important factors that significantly affected RNA production (confidence levels above 95%). These factors were further optimized using a central composite design (CCD) and RSM. The optimum values for the critical components were as follows: molasses 47.21 g/L: ZnSO4 0.048 g/L; H3PO4 1.19 g/L. Under optimal conditions, RNA production was 2.56 g/L, which was in excellent agreement with the predicted value (2.561 g/L), and led to a 2.1-fold increase compare with that using the original medium in RNA production.

A review—biosynthesis of optically pure ethyl (S)-4-chloro-3-hydroxybutanoate ester: recent advances and future perspectives

Co-reporter:Qi Ye;Pingkai Ouyang

Applied Microbiology and Biotechnology 2011 Volume 89( Issue 3) pp:513-522

Publication Date(Web):2011 February

DOI:10.1007/s00253-010-2942-3

Ethyl (S)-4-chloro-3-hydroxybutanoate ester ((S)-CHBE) is a precursor of enantiopure intermediates used for the production of chiral drugs, including the cholesterol-lowering 3-hydroxy-3-methyl-glutaryl CoA reductase inhibitors (statins). The asymmetric reduction of ethyl 4-chloro-3-oxobutanoate ester (COBE) to (S)-CHBE by biocatalysis has several positive attributes, including low cost, mild reaction conditions, high yield, and a high level of enantioselectivity. During genome database mining of the yeast Pichia stipitis, our group found two novel carbonyl reductases (PsCRI and PsCRII) that have a promising future for the industrial production of (S)-CHBE with >99% enantiomeric excess. This review covers the main process of biosynthesis of (S)-CHBE: screening of microorganisms that catalyze the reduction of COBE to (S)-CHBE (I); gene cloning, expression, and characterization of carbonyl reductases for the production of (S)-CHBE in Escherichia coli (II); development of cofactor generation systems for regenerating cofactors (III); and biocatalysis of COBE to (S)-CHBE by recombinant E. coli (IV).

Preparation, characterization, and drug-release properties of pH/temperature-responsive poly(N-isopropylacrylamide)/chitosan semi-IPN hydrogel particles

Co-reporter:Xiaochun Chen;He Song;Ting Fang;Jianxin Bai;Jian Xiong

Journal of Applied Polymer Science 2010 Volume 116( Issue 3) pp:1342-1347

Publication Date(Web):

DOI:10.1002/app.31583

Abstract

Novel poly(N-isopropylacrylamide) (PNIPAAm)/chitosan (CS) semi-interpenetrating polymer network hydrogel particles were prepared by inverse suspension polymerization. The prepared particles were sensitive to both temperature and pH, and they had good reversibility in solution at different temperatures and pH values. The swelling ratios of PNIPAAm/CS hydrogel particles decreased slightly with the addition of CS, which did not shift the lower critical solution temperature. The drug-release behavior of the particles was investigated using cyclic adenosine 3′,5′-monophosphate (cAMP) as a model drug. The release of cAMP from the hydrogel particles was affected by temperature, pH, and the CS content in the particles. These results showed that semi-IPN hydrogel particles appeared to be of great promise in pH- and temperature-sensitive oral drug release. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2010

High-level production of heterologous proteins using untreated cane molasses and corn steep liquor in Escherichia coli medium

Co-reporter:Qi Ye;Ximu Li;Ming Yan;Hou Cao;Lin Xu

Applied Microbiology and Biotechnology 2010 Volume 87( Issue 2) pp:517-525

Publication Date(Web):2010 June

DOI:10.1007/s00253-010-2536-0

To develop an economical industrial medium, untreated cane molasses (UCM) was tested as a carbon source for fermentation culturing of Escherichia coli. To test the industrial application of this medium, we chose a strain co-expressing a carbonyl reductase (PsCR) and a glucose dehydrogenase (BmGDH). Although corn steep liquor (CSL) could be used as an inexpensive nitrogen source to replace peptone, yeast extract could not be replaced in E. coli media. In a volume of 40 ml per 1-l flask, a cell concentration of optical density (OD600) 15.1 and enzyme activities of 6.51 U/ml PsCR and 3.32 U/ml BmGDH were obtained in an optimized medium containing 25.66 g/l yeast extract, 3.88 g/l UCM, and 7.1% (v/v) CSL. When 3.88 g/l UCM was added to the medium at 6 h in a fed-batch process, the E. coli concentration increased to OD600 of 24, and expression of both PsCR and BmGDH were twofold higher than that of a batch process. Recombinant cells from batch or fed-batch cultures were assayed for recombinant enzyme activity by testing the reduction of ethyl 4-chloro-3-oxobutanoate to ethyl (S)-4-chloro-3-hydroxybutanoate (CHBE). Compared to cells from batch cultures, fed-batch cultured cells showed higher recombinant enzyme expression, producing 560 mM CHBE in the organic phase with a molar yield of 92% and an optical purity of the (S)-isomer of >99% enantiomeric excess.

Solubility of Adenosine 5′-Monophosphate in Different Solvents from (288.15 to 330.15) K

Co-reporter:Dong Wang, Zhen-Jiang Li and Han-Jie Ying

Journal of Chemical & Engineering Data 2010 Volume 55(Issue 2) pp:992-993

Publication Date(Web):November 9, 2009

DOI:10.1021/je900462z

The solubilities of adenosine 5′-monophosphate (AMP) in water, methanol, ethanol, and acetic acid between (288.15 and 330.15) K were measured using a laser monitoring observation technique. Results of these measurements were correlated with a modified Apelblat equation, which can be used as a useful model in the crystallization process of AMP.

Solubility of Cytidine 5′-Diphosphocholine Sodium in Water and Different Binary Mixtures from (278.15 to 298.15) K

Co-reporter:Zhenxing Du, Xiaoquan Huang, Hanjie Ying, Jian Xiong, Hao Lv and Xiqun Zhou

Journal of Chemical & Engineering Data 2010 Volume 55(Issue 2) pp:1000-1002

Publication Date(Web):July 28, 2009

DOI:10.1021/je900465r

Experimental solubility data were measured for cytidine 5′-diphosphocholine sodium dissolved in water and methanol + water, ethanol + water, and acetone + water from (278.15 to 298.15) K. All of the experiments were carried out under atmospheric pressure by an isothermal method. The solubility data were correlated by the modified Apelblat equation. Computation showed that the model fit the data well.

Solubility of Sucralose in Different Solvents from (283.15 to 333.15) K

Co-reporter:Xin Li, Zhenxing Du, Xiaoquan Huang, Wei Yuan and Hanjie Ying

Journal of Chemical & Engineering Data 2010 Volume 55(Issue 7) pp:2600-2602

Publication Date(Web):January 14, 2010

DOI:10.1021/je9008427

The solubility of sucralose in water, methanol, ethanol, and isopropyl alcohol from (283.15 to 333.15) K was measured by an isothermal method. The results show that the solubility of sucralose in the four solvents increases with the increasing temperature. The experimental solubility data were correlated by the modified Apelblat equation, and the calculated values were in good agreement with the experimental data.

Optimization of medium components for D-ribose production by transketolase-deficient Bacillus subtilis NJT-1507

Co-reporter:Ting Fang;Xiaochun Chen;Nan Li;He Song

Korean Journal of Chemical Engineering 2010 Volume 27( Issue 6) pp:1725-1729

Publication Date(Web):2010 November

DOI:10.1007/s11814-010-0288-2

Statistical experimental designs were used to optimize the composition of culture media for the production of D-ribose by Bacillus subtilis. A fractional factorial design 2(5-2) was used to determine medium components that significantly affected D-ribose production. The concentrations of glucose and (NH4)2SO4 were the significant factors. Central composite design and response surface methodology were then used to estimate the quadratic response surface and determine the factor levels for maximum production of D-ribose. Finally, the optimal medium composition was obtained (g/L): glucose, 172.75; (NH4)2SO4, 13.2; yeast powder, 4; corn steep liquor, 8 and MnSO4, 0.5. This optimization strategy increased D-ribose production from 73.21 g/L to 88.57 g/L, an increase of 22% compared with the original conditions. The D-ribose production yield to glucose concentration was also enhanced from 0.37 g/g to 0.52 g/g. Confirmatory experiments were also performed to demonstrate the accuracy of the model. Under the optimal medium using ammonia to control pH in a 5 L fermenter, the D-ribose yield was increased to 95.28 g/L after 3 days of cultivation at 37 °C.

Biocatalytic synthesis of (S)-4-chloro-3-hydroxybutanoate ethyl ester using a recombinant whole-cell catalyst

Co-reporter:Qi Ye;Hou Cao;Guanglou Zang;Lan Mi;Ming Yan

Applied Microbiology and Biotechnology 2010 Volume 88( Issue 6) pp:1277-1285

Publication Date(Web):2010 December

DOI:10.1007/s00253-010-2836-4

A cofactor regeneration system for enzymatic biosynthesis was constructed by coexpressing a carbonyl reductase from Pichia stipitis and a glucose dehydrogenase from Bacillus megaterium in Escherichia coli Rosetta (DE3) PlySs. Transformants containing the polycistronic plasmid pET-PII-SD2-AS1-B exhibited an activity of 13.5 U/mg protein with 4-chloro-3-oxobutanoate ethyl ester (COBE) as the substrate and an activity of 14.4 U/mg protein with glucose as the substrate; NAD(H) was the coenzyme in both cases. Asymmetric reduction of COBE to (S)-4-chloro-3-hydroxybutanoate ethyl ester [(S)-CHBE] with more than 99% enantiomeric excess was demonstrated by transformants. Furthermore, the paper made a comparison of crude enzyme catalysis and whole-cell catalysis in an aqueous monophasic system and a water/organic solvent biphasic system. In the water/n-butyl acetate system, the coexpression system produced 1,398 mM CHBE in the organic phase, which is the highest yield ever reported for CHBE production by NADH-dependent reductases from yeasts. In this case, the molar yield of CHBE was 90.7%, and the total turnover number, defined as moles (S)-CHBE formed per mole NAD+, was 13,980.

The mechanisms of citrate on regulating the distribution of carbon flux in the biosynthesis of uridine 5′-monophosphate by Saccharomyces cerevisiae

Co-reporter:Yong Chen;Shuya Li;Jian Xiong;Zhenjiang Li

Applied Microbiology and Biotechnology 2010 Volume 86( Issue 1) pp:75-81

Publication Date(Web):2010 March

DOI:10.1007/s00253-009-2287-y

A whole cell biocatalytic process for uridine 5′-monophosphate (UMP) production from orotic acid by Saccharomyces cerevisiae was developed. The concentration of UMP was increased by 23% when 1 g l−1 sodium citrate was fed into the broth. Effects of citrate addition on UMP production were investigated. Glucose-6-phosphate pool was elevated by onefold, while FBP and pyruvate were decreased by 42% and 40%, respectively. Organic acid pools such as acetate and succinate were averagely decreased by 30% and 49%. The results demonstrated that manipulation of citrate levels could be used as a novel tool to regulate the metabolic fluxes distribution among glycolysis, pentose phosphate pathway, and TCA cycle.

Enhanced uridine diphosphate N-acetylglucosamine production using whole-cell catalysis

Co-reporter:Hanjie Ying;Xiaochun Chen;Haiping Cao

Applied Microbiology and Biotechnology 2009 Volume 84( Issue 4) pp:677-683

Publication Date(Web):2009 September

DOI:10.1007/s00253-009-2016-6

Uridine diphosphate N-acetylglucosamine (UDPAG) can be produced by chemical, enzymatic, chemoenzymatic, and fermentative methods. In this study, we used whole-cell catalysis method to produce UDPAG for the first time by Saccharomyces cerevisiae. In order to increase the ATP utilization efficiency and UDPAG conversion yield, the response surface methodology was applied to optimize the whole-cell catalytic conditions for UDPAG production. Firstly, effects of uridine 5′-monophosphate (5′-UMP), glucosamine, vitamin B1, glycerol, magnesium chloride, potassium chloride, temperature, sodium dihydrogen phosphate, sodium acetate, fructose, and pH on UDPAG production were evaluated by a fractional factorial design. Results showed that UDPAG production was mainly affected by sodium dihydrogen phosphate, temperature, and vitamin B1. Then, the concentrations of sodium dihydrogen phosphate and vitamin B1 and temperature were further investigated with a central composite design and response surface analysis. The cultivation conditions to obtain the optimal UDPAG production were determined: sodium dihydrogen phosphate, 31.2 g/L; temperature, 29°C, and vitamin B1, 0.026 g/L. This optimization strategy led to an enhancement of UDPAG production from 2.51 to 4.25 g/L, yield from 44.6% to 75.6% based on the initial 5′-UMP concentration, and ATP utilization efficiency from 7.43% to 12.6%.

A novel carbonyl reductase from Pichia stipitis for the production of ethyl (S)-4-chloro-3-hydroxybutanoate

Co-reporter:Qi Ye;Ming Yan;Lin Xu;Hou Cao;Zhenjiang Li;Yong Chen

Biotechnology Letters 2009 Volume 31( Issue 4) pp:537-542

Publication Date(Web):2009 April

DOI:10.1007/s10529-008-9907-y

An NADPH-dependent carbonyl reductase (PsCR) gene from Pichia stipitis was cloned. It contains an open reading frame of 849 bp encoding 283 amino acids whose sequence had less than 60% identity to known reductases that produce ethyl (S)-4-chloro-3-hydroxybutanoates (S-CHBE). When expressed in Escherichia coli, the recombinant PsCR exhibited an activity of 27 U/mg using ethyl 4-chloro-3-oxobutanoate (COBE) as a substrate. Reduction of COBE to (S)-CHBE by transformants in an aqueous mono-phase system for 18 h, gave a molar yield of 94% and an optical purity of the (S)-isomer of more than 99% enantiomeric excess.

Kinetics of Adsorption of Thymopentin on a Gel-Type Strong Cation-Exchange Resin

Co-reporter:Xin Li;Lei Zhang;Yonghui Chang;Shubao Shen

Chromatographia 2007 Volume 66( Issue 3-4) pp:231-235

Publication Date(Web):2007 August

DOI:10.1365/s10337-007-0291-x

The ion-exchange kinetics have been determined for adsorption of thymopentin on a gel-type sulfonated styrene–divinylbenzene resin converted to the ammonium form. Batch equilibrium and kinetic experiments were performed in chloride ion solutions of different concentration. Equilibrium data revealed isotherms were a good fit to the constant separation factor isotherm. Because of the high capacity and low cost of the resin its use for uptake of thymopentin was economically feasible. Kinetic data were compared with the predictions from the Nernst–Planck and Fick models. The intraparticle and effective diffusivity of thymopentin were obtained from these models.

Reversible, selective immobilization of nuclease P1 from a crude enzyme solution on a weak base anion resin activated by polyethylenimine

Co-reporter:Bingbing Li, Yong Chen, Zhi Cao, Huanqing Niu, Dong Liu, Ying He, Xiaochun Chen, Jinglan Wu, Jingjing Xie, Wei Zhuang, Hanjie Ying

Journal of Molecular Catalysis B: Enzymatic (March 2014) Volume 101() pp:92-100

Publication Date(Web):1 March 2014

DOI:10.1016/j.molcatb.2013.12.021

•Reversible immobilization of Nuclease P1 on PEI-activated resin.•Immobilization of crude nuclease P1 by selection adsorption.•Immobilization lowered the activation energy of enzyme.•The support could be regenerated after desorption of the inactivated enzyme.•The Immobilized enzyme enhanced the enzyme selectivity.The reversible immobilization of crude nuclease P1 fermented by Penicllium citrinum and used without further purification was performed via adsorption on a weak base anion resin activated by polyethylenimine (PEI). The immobilization conditions, including PEI concentration, the amount of support, the immobilization time, and the reusability of the PEI-activated resin were investigated. The results have shown that the PEI-activated resin has the capability of selectively adsorbing nuclease P1. Subsequently, the functional properties of the immobilized nuclease P1 were studied and compared to those of the free enzyme. The apparent Km value for immobilized nuclease P1 on the activated resin (15.31 g L−1) was about 4.41-fold higher than that of the free enzyme (3.47 g L−1), and the apparent Vmax value of the immobilized enzyme (530 U g−1) was about 3.9-fold less than that of the free enzyme (2082 U mL−1). The optimum temperature was observed to be 70 °C, 15 °C higher than that of the free enzyme. The optimum pH was the same for both free and immobilized nuclease P1 (pH 5.0). The apparent activation energies (Ea) of the free and immobilized nuclease P1 were 163.09 kJ mol−1 and 156.32 kJ mol−1, respectively, implying that the catalytic efficiency of the immobilized enzyme was restricted by mass-transfer rather than kinetic limitations.Download full-size image

Metabolic flux analysis of Arthrobacter sp. CGMCC 3584 for cAMP production based on 13C tracer experiments and gas chromatography–mass spectrometry

Co-reporter:Huanqing Niu, Yong Chen, Shiwei Yao, Lixia Liu, Chen Yang, Bingbing Li, Dong Liu, Jingjing Xie, Xiaochun Chen, Jinglan Wu, Hanjie Ying

Journal of Biotechnology (December 2013) Volume 168(Issue 4) pp:355-361

Publication Date(Web):1 December 2013

DOI:10.1016/j.jbiotec.2013.09.005

•13C metabolic flux analysis was performed for Arthrobacter sp. CGMCC 3584.•The stoichiometric growth model of Arthrobacter sp. CGMCC 3584 was constructed.•The central carbon metabolism of Arthrobacter sp. CGMCC 3584 was determined.•The fluxes through the PP pathway increased with hypoxanthine and NaF addition.Arthrobacter sp. CGMCC 3584 are able to produce cAMP from glucose by the purine synthesis pathway via de novo or salvage biosynthesis. In order to gain an improved understanding of its metabolism, 13C-labeling experiment and gas chromatography–mass spectrometry (GC–MS) analysis were employed to determine the metabolic network structure and estimate the intracellular fluxes. GC–MS analysis helps to reflect the activity of the intracellular pathways and reactions. The metabolic network mainly contains glycolytic and pentose phosphate pathways, the tricarboxylic acid cycle, and the inactive glyoxylate shunt. Hypoxanthine as a precursor of cAMP and sodium fluoride as an inhibitor of glycolysis were found to increase the cAMP production, as well as the flux through the PP pathway. The effects of adding hypoxanthine and sodium fluoride are discussed based on the enzyme assays and metabolic flux analysis. In conclusion, our results provide quantitative insights into how cells manipulate the metabolic network under different culture conditions and this may be of value in metabolic regulation for desirable production.