Methanol Feed Rate Research Articles

Development of pure hydrogen generation system based on methanol steam reforming and Pd membrane

Methanol steam reforming (MSR) has been used for in-situ hydrogen production, while hydrogen purification remains a technical challenge to limit the applications of using hydrogen for fuel cells. This study presents a pure hydrogen generation system (PHGS) by integrating MSR and hydrogen purification modules, this system enables low-temperature reforming and high-temperature purification to achieve high yield and recovery of hydrogen. To investigate the combined effects of feedstock flow rates and pressure on MSR and purification, the performance of each module and overall PHGS system were tested. The results showed that PHGS can stably produce high purity hydrogen (>99.99%) under different conditions, the synergistic effects of two modules lead to hydrogen yield exceeding the numerical sum of individual modules. At methanol feed rate of 1.00 g/min and system pressure of 5.0 bar, the pure hydrogen flow rate, CO content, pure hydrogen yield, hydrogen recovery and thermal power output were tested about 1.70 ± 0.01 L/min, 2.4 ± 0.1 ppm, 81.1 ± 0.5%, 87.6 ± 0.5% and 350 W. The PHGS can deliver up to 500 W of thermal power and is suitable for small range-extended electric vehicles. Our work on the developed PHGS can provide a significant reference for future practical applications of systems combining MSR and membrane purification.

Esterification and transesterification of palm fatty acid distillate in chitosan membrane reactor

Biodiesel from palm fatty acid distillate (PFAD) with FFAs of 87.1 ± 1.3 wt% was studied in a 2-litres chitosan membrane reactor. The reactor used in this study comprised of three parts; i.e., upstream side for reaction, membrane cell and downstream side connecting with a vacuum pump. The feed was operated under esterification reaction followed by transesterification reaction without intermediate washing. Flat sheet symmetric dense uncrosslinked chitosan membrane was used preferably in the laboratory 2-litres reactor. Operating the reactor under the pervaporation process, the optimum esterification condition was at 1:15 of PFAD to methanol ratio, 2.0 wt% H2SO4 and 20 ml/min methanol feed rate for 120 min. The FFA and methyl ester contents in the esterification product were 1.09 ± 0.03 and 85.9 ± 0.1 wt%, respectively. On the transesterification condition at 1:6 of PFAD to methanol ratio, 1.0 wt% NaOH and 20 ml/min methanol feed rate for 60 min, the methyl ester was increased to 97.9 ± 0.0 wt%. On performance and potential application, an 8-litres reactor installing a set of membrane modules was studied. Several membrane modules with flat sheet reinforced crosslinked composite chitosan membranes were installed in the pilot 8-litres reactor. Together with operating the reactor under the pervaporation process, the esterification was operated at 1:15 of PFAD to methanol ratio and 1.5 wt% H2SO4 for 180 min followed by the transesterification reaction at 1:6 of PFAD to methanol ratio and 1.0 wt% NaOH for 60 min without intermediate washing. The methyl ester yield of 96.6 ± 0.02 wt% was obtained.

Особенности каталитической трансформации метанола в углеводороды в условиях микроструктурированных потоков

The methanol into hydrocarbons transformation is a complex catalytic reaction accompanied by the formation of a wide range of hydrocarbons and proceeding on the surface of acid sites of various zeolites. Zeolite H-ZSM-5 considered to be most often used catalyst for this process. H-ZSM-5 is a highly dispersed material with a crystal diameter of 1–20 microns, which complicates its direct use in reactors with a fixed catalyst bed due to the high hydraulic pressure drop of the catalytic bed. Traditionally in industry, this issue is solved by using complex reactor systems with a fluidized bed, which is justified for large-scale production. In small and medium-size plants, the use of fluidized bed systems is not economically feasible. One of the possible solutions to this problem is the use of a monolithic catalyst with a supported layer of H-ZSM-5 zeolite. This article presents a study of the catalytic activity of a zeolite-containing microstructured monolith in methanol into hydrocarbons transformation. The monolith was synthesized by pressing a zeolite-containing mass followed by drying, calcining, and secondary growth of the zeolite on the monolith surface. A sample of a monolith with an average channel diameter of 0.5, 1.0, 1.5, 2.0 mm were synthesized this way. Samples of the microstructured catalyst were tested at varying temperatures from 250 to 450 °C and at varying the specific methanol feed rate from 0.65 to 2.3 kg (MeOH)/(kg (Cat) h). For this purpose, the monolithic catalyst was placed in a reactor for testing microstructured catalysts, which consisted of a pump, a temperature controller, a catalytic reactor, a condenser, a separating funnel, and a chromatograph. Varying the conditions showed that for the preferential production of gaseous C1–C4 hydrocarbons, it is advisable to carry out the reaction under the following conditions: the average diameter of the catalyst channels is 2 mm, the reaction temperature is 350 °C, the methanol feed rate is 1.65 kg (MeOH)/(kg (Cat) h). For the predominant formation of liquid hydrocarbons of the C5–C8 fraction, it is advisable to carry out the transformation of methanol into hydrocarbons under the following conditions: the average diameter of the catalyst channels is 1 mm, the reaction temperature is 350 °C, the methanol feed rate is 0.65 kg (MeOH) / (kg (Cat) h). For the predominant formation of liquid hydrocarbons of the C9–C12 fraction, it is advisable to carry out the transformation of methanol into hydrocarbons under the following conditions: the average diameter of the catalyst channels is 0.5 mm, the reaction temperature is 350 °C, and the methanol feed rate is 0.65 kg (MeOH) / (kg (Cat) h).

HBsAg Production in Methanol Controlled <i>P. pastoris</i> GS115 Mut<sup>S</sup> Bioreactor Process

When producing recombinant proteins with Pichia pastoris, cultivation parameters, such as induction temperature, dissolved oxygen level and residual methanol concentration play a crucial role in product biosynthesis and subsequent purification, therefore to maximize protein yields, the optimization of these parameters is imperative. Two different Pichia pastoris cultivation strategies for HBsAg VLP production in a 5 L stirred-tank bioreactor and the influence of different cultivation parameters on product yield were investigated. Residual methanol concentrations were controlled at low (>0.01 g/L), medium (1.5-2.0 g/L) and high (5.0-6.0 g/L) levels using a PI-based feed rate control algorithm based on the online methanol sensor signal. Product was purified using a novel and rapid purification method including steps of ammonium sulfate precipitation, size-exclusion chromatography and hydrophobic interaction chromatography. Employing an in-situ methanol sensor probe, the PI-based methanol feed rate control algorithm provided residual methanol concentration control with an average deviation of ±0.4 g/L from set-point value. Employing a cultivation protocol with an increased methanol concentration controlled at 6.0 g/L and a reduced DO level below 10 %, resulting in a final dry cell biomass concentration of 140 g/L and purified HBsAg VLPs yield of 186 mg/L. Developed purification method proved advantageous to other described methods, as it did not include time consuming extraction and centrifugation steps.

Synthesis and Characterization of Fe/SBA-15 Heterogeneous Catalysts for Methyl Acetate Production

In the present study, the synthesis of SBA-15 (Santa Barbara Amorphous) support material(SPm) was initially carried out with the method determined according to the literature research. The catalytic property of SBA-15 was imparted by the hydrothermal method (HtM), different from the traditionally applied impregnation method. In this method, the active substance (iron III oxide; Fe2O3) was added to the solution during the synthesis of the SPm. The amount of active substance in the synthesized catalysts was calculated based on the mass ratios of silicon (Si) in SBA-15 and iron (Fe) in Fe2O3. The active substance ratios in the catalysts were determined as 10, 25% Fe/Si. The activities of Fe/SBA-15 catalysts were tested in methyl acetate production (MAP). MAP reaction experiments were carried out in the presence of 0.4g catalyst (Fe/SBA-15), 373K, in autogenous pressure and 2/1 Acetic acid(AA)/methanol(MeOH) feed rate for 48 hours. The effects of active substance loading rate on AA conversion were determined. The structural properties of Fe/SBA-15 catalysts were investigated by XRD, BET, FTIR/DRIFT, SEM/EDX, and MAPPING analysis methods.

Transesterification Using Ultrasonic Spray of Triolein Containing CaO Particles into Methanol Vapor in a 3-Phase Reactor

Ultrasonic spraying was used in a three-phase reactor to produce small droplets of triolein mixed with CaO as a solid catalyst at temperatures above the boiling point of methanol for enhancement of the transesterification of triolein. Droplets fell in the methanol countercurrent flow and were collected at the bottom of the reactor, followed by circulation to the ultrasonic spray system. The experimental parameters included triolein flow rates of 2.5–9.0 mL/min, reaction temperatures of 70–100 °C, and catalyst contents of 1.0–7.0 wt%. The methanol feed rate was set to be constant. The results suggested that the enhancement was successful after using the three-phase reactor by generating a high contact surface area for the droplets, which was a key factor for determining the performance. Comparing the results with conventional transesterification in the liquid phase using the same CaO at 60 °C, the three-phase reactor produced a methyl ester yield 2–5% higher during the 60 min trial period. However, the yield became lower after 60 min because the mass transfer of methanol to the droplets was limited. The transesterification kinetics were estimated based on the experimental data—assuming a first-order reaction—and the results indicated a range of the rate constant, an apparent activation energy, and a pre-exponential factor of 1.21–3.70 × 10−2 min−1, 36.1 kJ mol−1, and 64.9 min−1, respectively, suggesting that the three-phase reactor was effective for fast transesterification at the initial stage.

Feasible production of hydrogen from methanol reforming through single stage DC microplasma reactor

Plasma-assisted methanol reforming is an effective technology to produce hydrogen for various clean energy applications. In this study, hydrogen was produced from methanol reforming in a unique single stage microplasma reactor. Microplasma was produced between the capillary stainless steel tube electrodes by using high voltage direct current (DC) power supply. Blend of methanol and water was supplied to the microplasma reactor in a controlled flow rate using nitrogen as carrier gas. The effects of applied input power to the discharge and methanol feed rate on the performance of the plasma methanol decomposition were investigated. The experimental results showed that increasing the applied input power expressively increased the methanol conversion and hydrogen energy yield. In contrast, the increased feed rate significantly decreased the methanol conversion efficiency though it enriched the hydrogen energy yield. Under selective conditions, hydrogen energy yield of 24.14 g kW[Formula: see text] h[Formula: see text] was achieved with the conversion efficiency of 71% and 50% selectivity for H2, which is comparatively better than many of plasma-assisted methanol reforming processes. This investigation reveals that methanol reforming through a single stage microplasma reactor has the ability to produce hydrogen efficiently without coke formation at room-temperature and atmospheric pressure.

Control of specific growth rate for the enhanced production of human interferon α2b in glycoengineered Pichia pastoris: process analytical technology guided approach

AbstractBACKGROUNDProcess variability in bioprocess systems involving genetically engineered strains is a common bottleneck and a real‐time insight of the on‐going process is crucial to achieve the desired product and its quality. In this study, a process analytical technology (PAT) platform was developed for the monitoring and control of specific growth rate during methanol induction phase (μmet) of glycoengineered Pichia pastoris fermentation for human interferon alpha 2b (huIFNα2b) production.RESULTSThe PAT guided approach involves real‐time monitoring of capacitance (ΔC) facilitating online estimation of specific growth rate (μest), which serves as a process input during controller application. Fed‐batch experiments using pulsed‐feeding of methanol at different dosage (20 g and 30 g) did not significantly influence μmet. Exponential methanol feeding was achieved using a modified proportional, integral and derivative (PID) controller for different predefined specific growth rate set point (μsp) values, namely 0.015, 0.03, 0.04 and 0.06 h−1. Exponential feeding strategy during the induction phase resulted in two crucial outcomes: (i) controlled methanol feeding rate (regulated by the developed PID controller) balanced the methanol consumption rate (qs, met) of the P. pastoris; (ii) significant improvement in huIFNα2b titer (1483 mg L−1) and specific productivity(>0.4 mg g−1. h) was achieved by the robust control of μmet at optimal (0.04 h−1) value. The purified huIFNα2b was found to exhibit antiproliferative effect against human breast cancer cell lines.CONCLUSIONSEfficient control of μmet at a very low narrow range was achieved with a long‐term controller stability (>10 h) and the highest titer reported to date for huIFNα2b (at optimal μmet) in the yeast expression platform. © 2019 Society of Chemical Industry

Modified Zeolite in Methanol to Hydrocarbons Transformation

The work is devoted to the modification of H-ZSM-5 samples with sodium hydroxide and oxalic acid to provide the formation of the mesoporous structure of the zeolite and to obtain optimal Si/Al ratio. Moreover, zeolite active sites were modified with Co, Ni, Fe in order to decrease polyaromatic substances formation during methanol transformation into hydrocarbons. The testing of the samples was performed at 350 oC in a flow reactor, the methanol feed rate was maintained at 0.16 g(methanol)/(g(Cat)*h). Dimethyl ether transformation rate into hydrocarbons was found to be 0.055-0.1 kg(DME)/(kg(Cat)*h) for investigated catalysts. Alkali treatment of H-ZSM-5 samples results in the formation of a mesoporous structure with a mean pore diameter of 5 nm. Formation of additional mesopores results in some increase of dimethyl ether to hydrocarbons transformation rate up to 0.09 kg(DME)/(kg(Cat)*h). Zeolite treatment with oxalic acid results in decreasing of Si/Al ratio from 0.76 to 0.41 that case some decrease of dimethyl ether transformation rate from 0.07 kg(DME)/(kg(Cat)*h) to 0.05 kg(DME)/(kg(Cat)*h), however considerable decrease of polyaromatic hydrocarbons concentration in reaction media was achieved for sample with lowest Si/Al ratio. Modification of H-ZSM-5 with Co, Fe, Ni results in some increase of dimethyl ether transformation rate up to 0.07-0.1 kg(DME)/(kg(Cat)*h) and a considerable decrease of polyaromatic hydrocarbons concentration in reaction media due to active sites modification.The ZSM-5-Ni-0.001M sample showed highest activity 0.1 kg(DME)/(kg(Cat)*h).

Application of recurrent neural network for online prediction of cell density of recombinant Pichia pastoris producing HBsAg

Artificial neural networking (ANN) seems to be a promising soft sensor for implementing current approaches of quality by design (QbD) and process analytical technologies (PAT) in the biopharmaceutical industry. In this study, we aimed to implement best-fitted ANN architecture for online prediction of the biomass amount of recombinant Pichia pastoris (P. pastoris) – expressing intracellular hepatitis B surface antigen (HBsAg) – during the fed-batch fermentation process using methanol as a sole carbon source. For this purpose, at the induction phase of methanol fed-batch fermentation, carbon evolution rate (CER), dissolved oxygen (DO), and methanol feed rate were selected as input vectors and total wet cell weight (WCW) was considered as output vector for the ANN. The obtained results indicated that after training recurrent ANN with data sets of four fed-batch runs, this toolbox could predict the WCW of the next fed-batch fermentation process at each specified time point with high accuracy. The R-squared and root-mean-square error between actual and predicted values were found to be 0.9985 and 13.73, respectively. This verified toolbox could have major importance in the biopharmaceutical industry since recombinant P. pastoris is widely used for the large-scale production of HBsAg.

PECULIARITIES OF SMALL STRAINED ALICYCLE COMPOUNDS FORMATION IN CATALYTIC TRANSFORMATION OF METHANOL OVER ZEOLITE H-ZSM-5

The article presents the results of strained hydrocarbons formation study during the catalytic transformation of methanol into hydrocarbons on zeolite H-ZSM-5. The formation of the following strained cyclic compounds was determined: 1,1-dimethylcyclopropane, 1,2-dimethyl-cyclopropane, 1,1,2-trimethylcyclopropane, 1,2,3-trimethylcyclopropane, 1,1,2,2-tetramethylcyclo-propane, 1,1,2 , 3-tetramethylcyclopropane. The non-stationary character of strained cyclic hydrocarbons formation with a pronounced hydrocarbons formation rate maximum and subsequent deactivation of the catalyst was found. The temperature effect on strained hydrocarbons yield was evaluated. Thus, with an increase in the process reaction temperature up to 400 °C, a maximum of strained hydrocarbons accumulation rate was achieved as 8-8.5 g(Hyd)/(kg(Cat)·h) on 350 h of reaction, and a further increase in the reaction temperature leads to a decrease in the strained hydrocarbons accumulation rate. The effect of the methanol feed rate on the strained hydrocarbons formation rate was also studied. An increase in the methanol feed rate from 0.02 ml/min to 0.16 ml/min results in increase in the strained hydrocarbons formation rate up to 37 g (Hyd)/(kg(Cat)·h). The article presents results of H-ZSM-5 physicochemical study used by ammonia chemisorption, nitrogen phisisorption, X-ray photoelectron spectroscopy. Physicochemical studies of catalyst samples after the methanol transformation process to form strained hydrocarbons showed a twofold decrease in the number of acid sites from 1.2 mmol(NH3)/g (sample) to 0.3 mmol (NH3)/g(sample) and a significant decrease in surface area of micropores from 294 m2/g for the initial sample to 16 m2/g for the sample after the reaction. The X-ray diffraction spectroscopy method showed that the composition of the catalysts H-ZSM-5 surface includes carbon, oxygen, silicon and aluminum. Carbon concentration was found to be 4.3 at.% on the surface of the initial catalyst. While the carbon concentration increases up to 14.1 at.% during the reaction. Also oxygen content on the catalysts surface decreases from 59.9 to 53.4 at%, silica concentration decreases from 35.5 to 32.1 at.%. The following indicates the formation of a carbon surface layer over the catalysts.

Reduced methanol input induces increased protein output by AOX1 promoter in a trans-acting elements engineered Pichia pastoris.

High oxygen consumption and heat release caused by methanol catabolism usually bring difficulties to industrial scale-up and cost for protein expression driven by methanol-induced AOX1 promoter in Pichia pastoris. Here, reduced methanol feeding levels were investigated for expression of insulin precursor in a trans-acting elements engineered P. pastoris strain MF1-IP. Insulin precursor expression level reached 6.69g/(L supernatant) at the methanol feeding rate of 6.67mL/(h·L broth), which was 59% higher than that in the wild-type strain WT-IP at the methanol feeding rate of 12mL/(h·L broth). Correspondingly, the insulin precursor expression level in fermentation broth and maximum specific insulin precursor production rate was 137 and 77% higher than the WT-IP, respectively. However, oxygen consumption and heat evolution were reduced, and the highest oxygen consumption rate and heat evolution rate of the MF1-IP were 18.0 and 37.7% lower than the WT-IP, respectively.

A Two-Dimensional Multiphysics Coupling Model of a Middle and Low Temperature Solar Receiver/Reactor for Methanol Decomposition

In this paper, the endothermic methanol decomposition reaction is used to obtain syngas by transforming middle and low temperature solar energy into chemical energy. A two-dimensional multiphysics coupling model of a middle and low temperature of 150~300 °C solar receiver/reactor was developed, which couples momentum equation in porous catalyst bed, the governing mass conservation with chemical reaction, and energy conservation incorporating conduction/convection/radiation heat transfer. The complex thermochemical conversion process of the middle and low temperature solar receiver/reactor (MLTSRR) system was analyzed. The numerical finite element method (FEM) model was validated by comparing it with the experimental data and a good agreement was obtained, revealing that the numerical FEM model is reliable. The characteristics of chemical reaction, coupled heat transfer, the components of reaction products, and the temperature fields in the receiver/reactor were also revealed and discussed. The effects of the annulus vacuum space and the glass tube on the performance of the solar receiver/reactor were further studied. It was revealed that when the direct normal irradiation increases from 200 W/m2 to 800 W/m2, the theoretical efficiency of solar energy transformed into chemical energy can reach 0.14–0.75. When the methanol feeding rate is 13 kg/h, the solar flux increases from 500 W/m2 to 1000 W/m2, methanol conversion can fall by 6.8–8.9% with air in the annulus, and methanol conversion can decrease by 21.8–28.9% when the glass is removed from the receiver/reactor.

Dimensional effect of graphite flow field channels of a direct methanol fuel cell under different operating conditions

Direct methanol fuel cells are a potential candidate to replace traditional power sources for portable applications. The flow field design, manufacture, and optimization are of great significance to the cell performance. When the scale of flow channels decreases to the level of submillimeter‐scale, it is favourable to the reactant and product management. This paper focuses on the effectiveness of using a multi‐tooth planing technique to create submillimeter‐scale parallel channels in a graphite sheet. Besides the structural parameters of flow channels, a series of operating parameters are experimentally investigated, including methanol concentration, methanol feed rate, oxygen feed rate, cathode backpressure, and environmental temperature. Results indicate that the prepared channels promote a higher cell performance than the traditional design with a larger scale. It is beneficial to both the anode and cathode performances, but it has a more prominent effect at the anode. The methanol concentration of 4 mol/L yields the best performance. Using a relatively lower methanol feed rate below 0.5 mL/min has a more obvious effect on the fuel cell. The cell performance is insensitive to the change of cathode oxygen feed rate and backpressure especially when the oxygen can be sufficiently supplied. In this case we can use lower levels of oxygen feed rate and cathode backpressure. The cell temperatures and influence of environmental temperature are also discussed.

Experimental Study on the Reaction Conditions of a Methanol Steam Reforming Process

Abstract The effects of various reaction conditions on a methanol steam reforming (MSR) process were analyzed in this paper by adopting design of experiment (DOE) techniques. The control factors considered are the length and the temperature of the catalyst bed, the steam-to-carbon ratio and the space velocity of the feedstocks, and the catalyst loading within the MSR reactor. The performance indices (quality characteristics) to be analyzed are the methanol conversion, the hydrogen production rate, the CO concentration in the produced mixture, and the ratios of the hydrogen production rate to the methanol feed rate and the catalyst loading, respectively. The major objective of the study is to analyze the holistic influences and the individual significance of the aforementioned reaction conditions on the defined MSR performance indices. Results suggest that temperature is the most significant factor that positively correlates the defined performance indices. The performance of a MSR reactor can be enhanced by raising the reacting temperature. Nonetheless, the CO concentration in the produced mixture is also increased. According to the present work, the effects of the reaction conditions on a MSR process can be analyzed simultaneously meanwhile the optimal combination of the concerned control factors can be obtained.

RSM optimization of HSA/IL1Ra in Pichia pastoris overexpression strain and study of its in vivo activity in reducing hyperglycemia of GK rats.

Human serum albumin (HSA) and interleukin-1 receptor antagonist (IL1Ra) fusion protein is a potential long-acting drug in the treatment of type 2 diabetes. Previously, the expression level of HSA/IL1Ra in Pichia pastoris was successfully improved by increasing the gene copy number and coexpression with chaperone (protein disulfide isomerase) in our laboratory. However, the overexpression strain resulted in low production of high- cell-density fermentation. In this study, the culture medium was optimized in both flask and fermenter, and the optimum culture medium notably increased the productivity and stability of HSA/IL1Ra. To further improve the expression, response surface methodology was used to further optimize the culture condition through modeling three selected parameters (induction pH, induction temperature [T], and maximum methanol feed rate [Vm ]). The maximum yield of HSA/IL1Ra reached 1.1 g/L (10-fold higher than original fermentation condition) under the optimized culture condition (pH 7.0, T = 29℃ and Vm = 4.82 mL/L/H) in a 5-L fermenter. In addition, the degradation position of HSA/IL1Ra during fermentation was determined to be K571, serving as a potential target for genetic modification strategies to reduce the degradation. Finally, the in vivo activity study showed that HSA/IL1Ra maintained the therapeutic effect of IL1Ra in type 2 diabetes model rats meanwhile reducing the frequency of administration.

Dynamic relationship between the CO2 gas bubble behavior and the pressure drop characteristics in the anode flow field of an active liquid-feed direct methanol fuel cell

Gas management is one of the bottlenecks hindering the development of direct methanol fuel cells (DMFCs). This work investigates the effects of anode serpentine flow field design on the cell performance, gas bubble behavior and pressure drop characteristics of an active liquid-feed DMFC. Three types of flow field setup based on the traditional right-angle serpentine flow field (RASFF), rounded-corner serpentine flow field (RCSFF), and step-wise broadening serpentine flow field (SBSFF) are tested. The results show that the fillet corner of the RCSFF and broadening channels of the SBSFF contribute to the CO2 emission and uniform distribution of reactants, respectively. The use of RCSFF and SBSFF yields a higher cell performance than that of RASFF under most of our testing conditions. There exists a strong relationship between the gas bubble behavior and the pressure drop characteristics. Gas slugs are conducive to reducing the pressure drop between the inlet and outlet on the anode side, whereas gas bubbles are counterproductive. The results also indicate that with increasing current density, the gas slugs gradually play a dominant role instead of gas bubbles in each flow field but the opposite is true when the methanol feed rate increases.

Post‐crystallization cooling behavior of aqueous supersaturated ammonium dihydrogen phosphate solutions containing acetone and methanol antisolvents added at different rates

Experimental results of post‐crystallization cooling behavior of aqueous supersaturated ammonium dihydrogen phosphate (ADP) solutions containing acetone and methanol antisolvents fed at different rate RA are described and discussed. The supersaturated solutions were contained in a double‐walled crystallizer connected to a thermostating arrangement maintaining the temperature at 30 °C. It was found that: (1) there is a steady decrease in temperature Ts of aqueous ADP solutions with increasing time t, which follows Newton's law of cooling, with constant K as characteristic of solution composition and its value for a system increases with an increase in RA, and (2) at a particular acetone and methanol feeding rate RA, the value of constant K for aqueous ADP solutions is much higher than that obtained from the data of decrease in the temperature Tm of pure water. The experimental results are discussed from consideration of a steady decrease in solution supersaturation with increasing t as a result of diffusion of ions/molecules involved in transporting heat from aqueous ADP solutions contained in the crystallizer to the thermostating system.

Visualization of two-phase flow and temperature characteristics of an active liquid-feed direct methanol fuel cell with diverse flow fields

Direct methanol fuel cell (DMFC) is a promising power source for portable applications. This study aims to reveal the two-phase flow characteristics of an active liquid-feed DMFC by using the visualization method. Different flow fields based on the serpentine, parallel and porous patterns and their effects on reactant and product managements are experimentally investigated. Results show that the performance of serpentine flow field is closely influenced by the change of methanol feed rate but the parallel pattern shows less sensitivity. The use of serpentine flow field enables the fuel cell to be operated with a higher methanol concentration. A higher methanol feed rate promotes removal of the produced gas bubbles. The oxygen feed rate has a negligible effect on the fuel cell with a cathode serpentine flow field but produces a more obvious performance difference in the case of parallel pattern. The visualization tests indicate that the use of a higher oxygen flow rate is helpful in water removal. The temperature characteristics of the anode serpentine channel are evaluated and how the temperature behaviors relate to different operating conditions are accordingly discussed.

Pichia pastoris Mut(S) strains are prone to misincorporation of O-methyl-L-homoserine at methionine residues when methanol is used as the sole carbon source.

BackgroundOver the last few decades the methylotrophic yeast Pichia pastoris has become a popular host for a wide range of products such as vaccines and therapeutic proteins. Several P. pastoris engineered strains and mutants have been developed to improve the performance of the expression system. Yield and quality of a recombinant product are important parameters to monitor during the host selection and development process but little information is published regarding quality differences of a product produced by different P. pastoris strains.ResultsWe compared titer and quality of several Nanobodies® produced in wild type and MutS strains. Titer in fed-batch fermentation was comparable between all strains for each Nanobody but a significant difference in quality was observed. Nanobodies expressed in MutS strains contained a product variant with a Δ−16 Da mass difference that was not observed in wild type strains. This variant showed substitution of methionine residues due to misincorporation of O-methyl-l-homoserine, also called methoxine. Methoxine is likely synthesized by the enzymatic action of O-acetyl homoserine sulfhydrylase and we confirmed that Nanobodies produced in the corresponding knock-out strain contained no methoxine variants. We could show the incorporation of methoxine during biosynthesis by its addition to the culture medium.ConclusionWe showed that misincorporation of methoxine occurs particularly in P. pastoris MutS strains. This reduction in product quality could outweigh the advantages of using Mut strains, such as lower oxygen and methanol demand, heat formation and in some cases improved expression. Methoxine incorporation in recombinant proteins is likely to occur when an excess of methanol is present during fermentation but can be avoided when the methanol feed rate protocol is carefully designed.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-016-0499-2) contains supplementary material, which is available to authorized users.