Production kinetics of β-carotene from Planococcus sp. TRC1 with concomitant bioconversion of industrial solid waste into crystalline cellulose rich biomass

Cost effective bioprocessing of nutraceuticals in present global scenario is a matter of concern. This study explored Paper mill sludge (PMS) and sugarcane bagasse (SCB) as inexpensive substrate for Planococcus sp. TRC1 mediated valuable β-carotene production and residual treated biomass as value added crystalline cellulose source simultaneously. Both biomass supported significant bacterial growth reaching highest yield 38.54 ± 1.4 mg/g on PMS (36 h) and 47.13 ± 1.9 mg/g (48 h) on SCB in solid state fermentation. Luedeking-Piret model revealed growth associated production with α and much lower β values of 5.18 and 0.24 for PMS and 4.5 and 0.165 for SCB. Cost analysis exhibited decrementation of pigment cost/mg by 84 % compared to synthetic media. Optimum production conditions were 30 °C temperature, pH 7, 10 % inoculum and initial moisture content 80 % (PMS) and 85 % (SCB). TLC (R_f = 0.9), HPLC (RT = 7.646) and lambda max (465 nm) confirmed pigment’s β-carotene nature with significant antioxidant and antimicrobial activity. It showed stability at varied temperature, pH and light conditions along with negligible phytotoxicity on Vigna radiata. Planococcus sp. TRC1 delignified PMS (41 %) and SCB (38 %) and FT-IR, FESEM and XRD suggested crystalline nature of residual cellulose rich fraction shedding light on a biorefinery approach for valorization of industrial solid wastes.     

Photoluminescence and piezoelectric behaviour of Y2Zr2O7 pyrochlore‐based multifunctional materials and the influence of Eu3+ and Sm3+

Y₂Zr₂O₇‐doped with Eu³⁺ and Sm³⁺ phosphors were prepared for the first time as multifunctional smart materials using a solid‐state reaction method at 1400^oC. Thermal behaviour, crystal structure, surface morphology, and elemental analysis were characterized using thermogravimetric (TG) and differential thermal (DTA) analyses, X‐ray diffraction (XRD) and scanning electron microscope equipped with energy‐dispersive X‐ray spectroscopy (SEM‐EDX). Experimental results revealed that both phosphors have a pyrochlore structure with a cubic crystal system. Photoluminescence properties were also measured and red emission was observed from Y_1.90Eu_0.10Zr₂O₇ and Y_1.90Sm_0.10Zr₂O₇ phosphors. Dielectric constant, loss tangent, piezoelectric charge constant, and Curie temperature of all the samples were determined using an LCR‐meter, d₃₃‐meter, and TG/DTA. Eu doping in Y₂Zr₂O₇ resulted in a high dielectric constant (9.61) and low loss tangent (1.67%) values, whereas high piezoelectric charge constant (0.68 pC/N) and high Curie temperature (820°C) could be obtained using Sm‐doped Y₂Zr₂O₇.     

Micro‐ and nano‐scale mineralogical characterization of Fe(II)‐oxidizing bacterial stalks

Neutrophilic, microaerobic Fe(II)‐oxidizing bacteria (FeOB) from marine and freshwater environments are known to generate twisted ribbon‐like organo‐mineral stalks. These structures, which are extracellularly precipitated, are susceptible to chemical influences in the environment once synthesized. In this paper, we characterize the minerals associated with freshwater FeOB stalks in order to evaluate key organo‐mineral mechanisms involved in biomineral formation. Micro‐Raman spectroscopy and Field Emission Scanning Electron Microscopy revealed that FeOB isolated from drinking water wells in Sweden produced stalks with ferrihydrite, lepidocrocite and goethite as main mineral components. Based on our observations made by micro‐Raman Spectroscopy, field emission scanning electron microscopy and scanning transmission electron microscope combined with electron energy‐loss spectroscopy, we propose a model that describes the crystal‐growth mechanism, the Fe‐oxidation state, and the mineralogical state of the stalks, as well as the biogenic contribution to these features. Our study suggests that the main crystal‐growth mechanism in stalks includes nanoparticle aggregation and dissolution/re‐precipitation reactions, which are dominant near the organic exopolymeric material produced by the microorganism and in the peripheral region of the stalk, respectively.     

外源水杨酸对NaCl胁迫下白榆幼苗光合作用及离子分配的影响

以1年生白榆幼苗为研究对象,设置0、0.5、1.0和2.0 mmol·L^-1水杨酸(SA)与0、50、100和150 mmol·L^-1 NaCl处理组合,考察盐胁迫下白榆幼苗生物量、光合色素含量、光合作用参数及根叶离子含量、分配、运输的情况,探讨外源SA对NaCl胁迫下白榆幼苗耐盐生理特征的影响。结果表明:(1)NaCl胁迫显著抑制了白榆幼苗的生长、光合色素含量及光合能力,并破坏了白榆体内离子平衡。(2)喷施外源SA使盐胁迫下白榆幼苗的干重和根冠比均不同程度升高,0.5和2.0 mmol·L^-1 SA不同程度提高了50和100 mmol·L^-1 NaCl处理组幼苗叶片光合色素含量。(3)0.5 mmol·L^-1 SA显著提升了50 mmol·L^-1 NaCl处理组白榆幼苗的净光合速率(Pn)、气孔导度(Gs)和蒸腾速率(Tr),1.0和2.0 mmol·L^-1 SA对150 mmol·L^-1 NaCl处理组幼苗净光合速率改善效果较好,外源SA对100 mmol·L^-1 NaCl处理组幼苗的光合作用参数无显著影响。(4)NaCl胁迫下,外源SA处理的白榆幼苗叶和根Na^+含量及Na^+/K^+、Na^+/Ca^2+和Na^+/Mg^2+显著降低,离子选择运输系数SK,Na、SCa,Na和SMg,Na升高,从而促进了幼苗K^+、Ca^2+和Mg^2+由根向叶片的转运;隶属函数分析发现对白榆幼苗叶和根中离子含量改善效果最好的SA浓度分别为1.0和2.0 mmol·L^-1。因此,适宜浓度的外源水杨酸能够有效改善NaCl胁迫下白榆幼苗的光合能力,有效调节白榆幼苗体内离子状态,从而增强白榆对NaCl胁迫的抗性。     

Model-based design and control of a small-scale integrated continuous end-to-end mAb platform

A continuous integrated bioprocess available from the earliest stages of process development allows for an easier, more efficient and faster development and characterization of an integrated process as well as production of small-scale drug candidates. The process presented in this article is a proof-of-concept of a continuous end-to-end monoclonal antibody production platform at a very small scale based on a 200 ml alternating tangential flow filtration perfusion bioreactor, integrated with the purification process with a model-based design and control. The downstream process, consisting of a periodic twin-column protein A capture, a virus inactivation, a CEX column and an AEX column, was compactly implemented in a single chromatography system, with a purification time of less than 4 hr. Monoclonal antibodies were produced for 17 days in a high cell density perfusion culture of CHO cells with titers up to 1.0 mg/ml. A digital twin of the downstream process was created by modelling all the chromatography steps. These models were used for real-time decision making by the implementation of control strategies to automatize and optimize the operation of the process. A consistent glycosylation pattern of the purified product was ensured by the steady state operation of the process. Regarding the removal of impurities, at least a 4-log reduction in the HCP levels was achieved. The recovery yield was up to 60%, and a maximum productivity of 0.8 mg/ml/day of purified product was obtained.     

Multivariate data analysis of growth medium trends affecting antibody glycosylation

Use of multivariate data analysis for the manufacturing of biologics has been increasing due to more widespread use of data-generating process analytical technologies (PAT) promoted by the US FDA. To generate a large dataset on which to apply these principles, we used an in-house model CHO DG44 cell line cultured in automated micro bioreactors alongside PAT with four commercial growth media focusing on antibody quality through N-glycosylation profiles. Using univariate analyses, we determined that different media resulted in diverse amounts of terminal galactosylation, high mannose glycoforms, and aglycosylation. Due to the amount of in-process data generated by PAT instrumentation, multivariate data analysis was necessary to ascertain which variables best modeled our glycan profile findings. Our principal component analysis revealed components that represent the development of glycoforms into terminally galacotosylated forms (G1F and G2F), and another that encompasses maturation out of high mannose glycoforms. The partial least squares model additionally incorporated metabolic values to link these processes to glycan outcomes, especially involving the consumption of glutamine. Overall, these approaches indicated a tradeoff between cellular productivity and product quality in terms of the glycosylation. This work illustrates the use of multivariate analytical approaches that can be applied to complex bioprocessing problems for identifying potential solutions.     

Determination of protein concentration in downstream biomanufacturing processes by in-line index of refraction

Protein concentration determination is a necessary in-process control for the downstream operations within biomanufacturing. As production transitions from batch mode to an integrated continuous bioprocess paradigm, there is a growing need to move protein concentration quantitation from off-line to in-line analysis. One solution to fulfill this process analytical technology need is an in-line index of refraction (IoR) sensor to measure protein concentration in real time. Here the performance of an IoR sensor is evaluated through a series of experiments to assess linear response, buffer matrix effects, dynamic range, sensor-to-sensor variability, and the limits of detection and quantitation. The performance of the sensor was also tested in two bioprocessing scenarios, ultrafiltration and capture chromatography. The implementation of this in-line IoR sensor for real-time protein concentration analysis and monitoring has the potential to improve continuous bioprocess manufacturing.     

Optimal Design of a Series of GSTR's Performing Reversible Reactions Catalyzed by Soluble Enzymes: A Theoretical Study

The condition for the minimum overall reactor volume of a given number of CSTR's in series is theoretically determined for a reversible, single reactant-single product (Uni-Uni) enzyme catalyzed reaction. The reactor network is assumed to operate in steady-state, isothermal conditions with a single phase and a constant activity of biocatalyst. The method is based on a mathematical analysis of the discrete substrate concentration profile along the CSTR's assuming complete micromixing. The algebraic equations describing the critical loci are obtained for the general case, the mathematical proof that these equations define a minimum is presented, and an exact solution arising from an asymptotic situation is found. An approximate analytical method of optimization based on the aforementioned critical behavior is reported and its validity and usefulness discussed. The formulae introduced can be used in more general situations as tools for getting the approximate range where the optimal overall volume of the series of CSTR's lies. Hence, the reasoning developed is important for the preliminary CSTR design and relevant in the initial steps of the more involved methods of numerical optimization. Finally, the enzymatic conversion of fumarate to L-malate is examined as a model system in order to assess the usefulness and applicability of the analysis developed.     

Effect of yeast cell disruption on ADH activity for redox reactions with in situ cofactor regeneration in a continuous solid/gas bioreactor

It has recently been demonstrated that dried cells of Saccharomyces cerevisiae were able to produce alcohols and aldehydes in a solid/gas reactor with in situ cofactor regeneration. Since diffusion of gaseous substrates may be limited by the membrane and cell wall, cell disruption by sonication was used to improve oxidoreduction with ethanol and butyraldehyde as substrates. Results showed that partial cell disruption enhances the maximum conversion yield with the best results obtained after 2 min of sonication. Beyond this time, the ADH activity decreased. Better stability was observed in the pellet obtained after centrifugation indicating the importance of cell environment for enzyme stability. Tests on purified mitochondria showed that the ADH activity in cells was mainly cytoplasmic. The addition of oxidized cofactor did not change either the activity or the stability of the catalyst in a gaseous medium. The effect of water activity was studied on material obtained after 2 min of disruption and a reduction of critical water activity needed for revealing enzymatic activity was observed. With increasing a_w, the enzyme was active at a_w=0.3 while a water activity of 0.4 was required before disruption. Nevertheless, the best compromise between activity and stability was obtained in both cases for a water activity of 0.57.     

Chinese plasma-derived products supply under the lot release management system in 2007–2011

In 2007, the Chinese State Food and Drug Administration (SFDA) implemented a management system for lot release of all plasma-derived products. Since then, there have been only a few systematic studies of the blood supply, which is a concern when considering the small amount of plasma collected per capita (approximately 3 L/1000 people). As a result, there may be a threat to the safety of the available blood supply. In this study, we examined the characteristics of the supply of Chinese plasma-derived products. We investigated the reports of lot-released biological products derived from all 8 national or regional regulatory authorities in China from 2007 to 2011. The market supply characteristics of Chinese plasma-derived products were analyzed by reviewing the changes in supply varieties, the batches of lot-released plasma-derived products and the actual supply. As a result, the national regulatory authorities can more accurately develop a specific understanding of the production and quality management information provided by Chinese plasma product manufacturers. The implementation of the lot release system further ensures the clinical validity of the plasma-derived products in China and improves the safety of using plasma-derived products. This work provides an assessment of the future Chinese market supply of plasma-derived products and can function as a theoretical basis for the establishment of hemovigilance.