Enhancing Soil Resilience: Bacterial Alginate Hydrogel vs. Algal Alginate in Mitigating Agricultural Challenges.

Erosion and tillage changes negatively the soil physical structure, which directly impacts agricultural systems and consequently food security. To mitigate these adverse modifications, different polymeric materials from synthetic and natural sources, have been used as soil conditioners to improve the hydro-mechanical behavior of affected soils. One of the most interesting and used natural polymers is the alginate hydrogel. Although commercially available alginate hydrogels are primarily sourced from algal, they can also be sourced from bacteria. The gelation capacity of these hydrogels is determined by their molecular properties, which, in turn, are influenced by the production conditions. Bacterial alginate hydrogel production offers the advantage of precise control over environmental conditions during cultivation and extraction, thereby maintaining and enhancing their molecular properties. This, in turn, results in higher molecular weight and improved gelation capacity. In this study, we compared the effects of bacterial alginate (BH) and algal alginate (AH) hydrogels over the mechanical, hydraulic, and structural behavior of coarse quartz sand as a model soil. Mechanically, it was observed that the treatment with the lowest concentration of bacteria alginate hydrogel (BH1) reached higher values of yield strength, Young's modulus (E), shear modulus (G) and strain energy (U) than those treatments with algal alginate hydrogel (AH). Furthermore, the increase in the aggregate stability could be associated with the improvement of mechanical parameters. On the other hand, a greater water retention capacity was observed in the BH treatments, as well as a greater decrease in hydraulic conductivity with respect to the AH and control treatments. All these changes could be explained by the formation of bridge-like structures between the sand particles and the hydrogel, and this alteration may result in a shift in the mechanical and wettability characteristics of the treated soils. Finally, our findings emphasize the superior impact of bacterial alginate hydrogel on enhancing the mechanical and hydraulic properties of coarse quartz sand compared to traditional algal alginate. Besides, the use of bacterial alginate hydrogel could be useful to counteract erosion and water scarcity scenarios in agricultural systems.

Relation of 3HV fraction and thermomechanical properties of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) produced by Azotobacter vinelandii OP.

Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) has attracted substantial attention as a promising material for industrial applications. In this study, different PHBV films with distinct 3-hydroxyvalerate (3HV) contents produced by Azotobacter vinelandii OP were evaluated. The 3HV fraction ranged from 18.6 to 36.7 mol%, and the number-average molecular weight (Mn) was between 238 and 434 kDa. In the bioreactor, a 3HV fraction (36.7 mol%) and an Mn value of 409 kDa were obtained with an oxygen transfer rate (OTR) of 12.5 mmol L-1 h-1. Thermal analysis measurements showed decreased melting (Tm) and glass transition (Tg) temperatures, and values with relatively high 3HV fractions indicated improved thermomechanical properties. The incorporation of the 3HV fraction in the PHBV chain improved the thermal stability of the films, reduced the polymer Tm, and affected the tensile strength. PHBV film with 36.7 mol% 3HV showed an increase in its tensile strength (51.8 MPa) and a decrease in its Tm (170.61 °C) compared with PHB. Finally, scanning electron microscopy (SEM) results revealed that the PHBV film with 32.8 mol% 3HV showed a degradation upon contact with soil, water, or soil bacteria, showing more porous surfaces after degradation. The latter phenomenon indicated that thermomechanical properties played an important role in biodegradation.

Production of poly (3-hydroxybutyrate) and extracellular polymeric substances from glycerol by the acidophile Acidiphilium cryptum.

Acidiphilium cryptum is an acidophilic, heterotrophic, and metallotolerant bacteria able to use dissolved oxygen or Fe(III) as an electron sink. The ability of this extremophile to accumulate poly(3-hydroxybutyrate) (PHB) and secrete extracellular polymeric substances (EPS) has also been reported. Hence, the aim of this work is to characterize the production of PHB and EPS by the wild strain DSM2389 using glycerol in shaken flasks and bioreactor. Results showed that maximum PHB accumulation (37-42% w/w) was obtained using glycerol concentrations of 9 and 15 g L-1, where maximum dry cell weight titers reached 3.6 and 3.9 g L-1, respectively. The culture in the bioreactor showed that PHB accumulation takes place under oxygen limitation, while the redox potential of the culture medium could be used for online monitoring of the PHB production. Recovered EPS was analyzed by Fourier-transform infrared spectroscopy and subjected to gas chromatography-mass spectrometry after cleavage and derivatization steps. These analyses showed the presence of sugars which were identified as mannose, rhamnose and glucose, in a proportion near to 3.2:2.3:1, respectively. Since glycerol had not been used in previous works, these findings suggest the potential of A. cryptum to produce biopolymers from this compound at a large scale with a low risk of microbial contamination due to the low pH of the fermentation process.

Efficient production of a polyhydroxyalkanoate by Azotobacter vinelandii OP using apple residues as promising feedstock.

Fruit residues are attractive substrates for the production of bacterial polyhydroxyalkanoates due to the high contents of fermentable sugars and the fast, simple, and efficient pretreatment methods required. In this study, apple residues, mainly apple peel, were used as the sole carbon source in cultures of the bacterium Azotobacter vinelandii OP to produce poly-3-hydroxybutyrate (P3HB). Conversion from the residue to total sugars was highly effective, achieving conversions of up to 65.4 % w w-1 when using 1 % v v-1 sulfuric acid and 58.3 % w w-1 in the absence of acid (only water). The cultures were evaluated at the shake-flask scale and in 3-L bioreactors using a defined medium under nitrogen starvation conditions. The results showed the production of up to 3.94 g L-1 P3HB in a bioreactor, reaching an accumulation of 67.3 % w w-1 when using apple residues. For the PHB obtained from the cultures with apple residues, a melting point of 179.99 °C and a maximum degradation temperature of 274.64 °C were calculated. A P3HB production strategy is shown using easily hydrolysable fruit residues to achieve production yields comparable to those obtained with pure sugars under similar cultivation conditions.

Extended batch cultures for poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) production by Azotobacter vinelandii OP growing at different aeration rates.

Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) is a polymer produced by Azotobacter vinelandii OP. In the bioreactor, PHBV production and its molar composition are affected by aeration rate. PHBV production by A. vinelandii OP was evaluated using extended batch cultures at different aeration rates, which determined different oxygen transfer rates (OTR) in the cultures. Under the conditions evaluated, PHBV with different 3-hydroxyvalerate (3HV) fractions were obtained. In the cultures with a low OTR (6.7 mmol L-1 h-1, at 0.3 vvm), a PHBV content of 38% w w-1 with 9.1 mol % 3HV was achieved. The maximum PHBV production (72% w w-1) was obtained at a high OTR (18.2 mmol L-1 h-1, at 1.0 vvm), both at 48 h. Thus, PHBV production increased in the bioreactor with an increased aeration rate, but not the 3HV fraction in the polymer chain. An OTR of 24.9 mmol L-1 h-1 (at 2.1 vvm) was the most suitable for improving the PHBV content (61% w w-1) and a high 3HV fraction of 20.8 mol % (at 48 h); and volumetric productivity (0.15 g L-1 h-1). The findings indicate that the extended batch culture at 2.1 vvm is the most adequate mode of cultivation to produce higher biomass and PHBV with a high 3HV fraction. Overall, the results have shown that the PHBV production and 3HV fraction could be affected by the aeration rate and the proposed approach could be applied to implement cultivation strategies to optimize PHBV production for different biotechnological applications.

Bacterial Alginate-Based Hydrogel Reduces Hydro-Mechanical Soil-Related Problems in Agriculture Facing Climate Change.

Agricultural systems are facing the negative impacts of erosion and water scarcity, directly impacting the hydro-mechanical behavior of soil aggregation. Several technologies have been proposed to reduce hydro-mechanical soil-related problems in agriculture. Biopolymer-based hydrogels have been reported to be a great tool to tackle these problems in soils. In this study, we investigated the hydro-mechanical behavior of different soils media treated with Ca-bacterial alginate hydrogel. We used an unconfined uniaxial compression test, aggregate stability test and hydraulic conductivity measurements to investigate the mechanical and hydraulic behavior of treated soils media. Our results from unconfined uniaxial compression test showed that yield stress (i.e., strength) increased in treated soils with higher kaolinite and water content (i.e., HCM3), compared with untreated coarse quartz sand (i.e., CM1). Furthermore, we found that temperature is an important factor in the gelation capacity of our hydrogel. At room temperature, HCM3 displayed the higher aggregate stability, almost 5.5-fold compared with treated coarse quartz sand (HCM1), while this differential response was not sustained at warm temperature. In general, the addition of different quantities of kaolinite decreased the saturated hydraulic conductivity for all treatments. Finally, bright field microscopy imaging represents the soil media matrix between sand and clay particles with Ca-bacterial alginate hydrogel that modify the hydro-mechanical behavior of different soils media. The results of this study could be helpful for the soil-related problems in agriculture facing the negative effects of climate change.

Obtaining Active Polylactide (PLA) and Polyhydroxybutyrate (PHB) Blends Based Bionanocomposites Modified with Graphene Oxide and Supercritical Carbon Dioxide (scCO2)-Assisted Cinnamaldehyde: Effect on Thermal-Mechanical, Disintegration and Mass Transport Properties.

Bionanocomposites based on Polylactide (PLA) and Polyhydroxybutyrate (PHB) blends were successfully obtained through a combined extrusion and impregnation process using supercritical CO2 (scCO2). Graphene oxide (GO) and cinnamaldehyde (Ci) were incorporated into the blends as nano-reinforcement and an active compound, respectively, separately, and simultaneously. From the results, cinnamaldehyde quantification values varied between 5.7% and 6.1% (w/w). When GO and Ci were incorporated, elongation percentage increased up to 16%, and, therefore, the mechanical properties were improved, with respect to neat PLA. The results indicated that the Ci diffusion through the blends and bionanocomposites was influenced by the nano-reinforcing incorporation. The disintegration capacity of the developed materials decreased with the incorporation of GO and PHB, up to 14 and 23 days of testing, respectively, without compromising the biodegradability characteristics of the final material.

Increases in alginate production and transcription levels of alginate lyase (alyA1) by control of the oxygen transfer rate in Azotobacter vinelandii cultures under diazotrophic conditions

BACKGROUND: Alginates are polysaccharides used in a wide range of industrial applications, with their functional properties depending on their molecular weight. In this study, alginate production and the expression of genes involved in polymerization and depolymerization in batch cultures of Azotobacter vinelandii were evaluated under controlled and noncontrolled oxygen transfer rate (OTR) conditions. RESULTS: Using an oxygen transfer rate (OTR) control system, a constant OTR (20.3 ± 1.3 mmol L 1 h 1 ) was maintained during cell growth and stationary phases. In cultures subjected to a controlled OTR, alginate concentrations were higher (5.5 ± 0.2 g L 1 ) than in cultures under noncontrolled OTR. The molecular weight of alginate decreased from 475 to 325 kDa at the beginning of the growth phase and remained constant until the end of the cultivation period. The expression level of alyA1, which encodes an alginate lyase, was more affected by OTR control than those of other genes involved in alginate biosynthesis. The decrease in alginate molecular weight can be explained by a higher relative expression level of alyA1 under the controlled OTR condition. CONCLUSIONS: This report describes the first time that alginate production and alginate lyase (alyA1) expression levels have been evaluated in A. vinelandii cultures subjected to a controlled OTR. The results show that automatic control of OTR may be a suitable strategy for improving alginate production while maintaining a constant molecular weight.

Molecular weight and guluronic/mannuronic ratio of alginate produced by Azotobacter vinelandii at two bioreactor scales under diazotrophic conditions.

Alginates can be used to elaborate hydrogels, and their properties depend on the molecular weight (MW) and the guluronic (G) and mannuronic (M) composition. In this study, the MW and G/M ratio were evaluated in cultures of Azotobacter vinelandii to 3 and 30 L scales at different oxygen transfer rates (OTRs) under diazotrophic conditions. An increase in the maximum OTR (OTRmax) improved the alginate production, reaching 3.3 ± 0.2 g L-1. In the cultures conducted to an OTR of 10.4 mmol L-1 h-1 (500 rpm), the G/M increased during the cell growth phase and decreased during the stationary phase; whereas, in the cultures at 19.2 mmol L-1 h-1 was constant throughout the cultivation. A higher alginate MW (520 ± 43 kDa) and G/M ratio (0.86 ± 0.01) were obtained in the cultures conducted at 10.4 mmol L-1 h-1. The OTR as a criterion to scale up alginate production allowed to replicate the concentration and the alginate production rate; however, it was not possible reproduce the MW and G/M ratio. Under a similar specific oxygen uptake rate (qO2) (approximately 65 mmol g-1 h-1) the alginate MW was similar (approximately 365 kDa) in both scales. The evidences revealed that the qO2 can be a parameter adequate to produce alginate MW similar in two bioreactor scales. Overall, the results have shown that the alginate composition could be affected by cellular respiration, and from a technological perspective the evidences contribute to the design process based on oxygen consumption to produce alginates defined.

Production of Poly-3-Hydroxybutyrate (P3HB) with Ultra-High Molecular Weight (UHMW) by Mutant Strains of Azotobacter vinelandii Under Microaerophilic Conditions.

Poly-3-hydroxybutyrate (P3HB) is a biopolymer, which presents characteristics similar to those of plastics derived from the petrochemical industry. The thermomechanical properties and biodegradability of P3HB are influenced by its molecular weight (MW). The aim of the present study was to evaluate the changes of the molecular weight of P3HB as a function of oxygen transfer rate (OTR) in the cultures using two strains of Azotobacter vinelandii, a wild-type strain OP, and PhbZ1 mutant with a P3HB depolymerase inactivated. Both strains were grown in a bioreactor under different OTR conditions. An inverse relationship was found between the average molecular weight of P3HB and the OTRmax, obtaining a polymer with a maximal MW (8000-10,000 kDa) from the cultures developed at OTRmax of 5 mmol L-1 h-1 using both strains, with respect to the cultures conducted at 8 and 11 mmol L-1 h-1, which produced a P3HB between 4000 and 5000 kDa. The increase in MW of P3HB was related to the activity of enzymes involved in the synthesis and depolymerization. Overall, our results show that it is possible to modulate the average molecular weight of P3HB by manipulating oxygen transfer conditions with both strains (OP and PhbZ1 mutant) of A. vinelandii.

Respiration in Azotobacter vinelandii and its relationship with the synthesis of biopolymers

Azotobacter vinelandii is a gram-negative soil bacterium that produces two biopolymers of biotechnological interest, alginate and poly(3-hydroxybutyrate), and it has been widely studied because of its capability to fix nitrogen even in the presence of oxygen. This bacterium is characterized by its high respiration rates, which are almost 10-fold higher than those of Escherichia coli and are a disadvantage for fermentation processes. On the other hand, several works have demonstrated that adequate control of the oxygen supply in A. vinelandii cultivations determines the yields and physicochemical characteristics of alginate and poly(3-hydroxybutyrate). Here, we summarize a review of the characteristics of A. vinelandii related to its respiration systems, as well as some of the most important findings on the oxygen consumption rates as a function of the cultivation parameters and biopolymer production.

Differential Gene Expression and Allele Frequency Changes Favour Adaptation of a Heterogeneous Yeast Population to Nitrogen-Limited Fermentations.

Alcoholic fermentation is fundamentally an adaptation process, in which the yeast Saccharomyces cerevisiae outperforms its competitors and takes over the fermentation process itself. Although wine yeast strains appear to be adapted to the stressful conditions of alcoholic fermentation, nitrogen limitations in grape must cause stuck or slow fermentations, generating significant economic losses for the wine industry. One way to discover the genetic bases that promote yeast adaptation to nitrogen-deficient environments are selection experiments, where a yeast population undergoes selection under conditions of nitrogen restriction for a number of generations, to then identify by sequencing the molecular characteristics that promote this adaptation. In this work, we carried out selection experiments in bioreactors imitating wine fermentation under nitrogen-limited fermentation conditions (SM60), using the heterogeneous SGRP-4X yeast population, to then sequence the transcriptome and the genome of the population at different time points of the selection process. The transcriptomic results showed an overexpression of genes from the NA strain (North American/YPS128), a wild, non-domesticated isolate. In addition, genome sequencing and allele frequency results allowed several QTLs to be mapped for adaptation to nitrogen-limited fermentation. Finally, we validated the ECM38 allele of NA strain as responsible for higher growth efficiency under nitrogen-limited conditions. Taken together, our results revealed a complex pattern of molecular signatures favouring adaptation of the yeast population to nitrogen-limited fermentations, including differential gene expression, allele frequency changes and loss of the mitochondrial genome. Finally, the results suggest that wild alleles from a non-domesticated isolate (NA) may have a relevant role in the adaptation to the assayed fermentation conditions, with the consequent potential of these alleles for the genetic improvement of wine yeast strains.

Biotransformation of poly(cis-1,4-isoprene) in a multiphase enzymatic reactor for continuous extraction of oligo-isoprenoid molecules.

Biotechnological processes for the partial degradation or transformation of poly(cis-1,4-isoprene) rubber have been investigated during recent decades with promising results. The use of the enzyme 'latex clearing protein' (Lcp) to transform the polymer into more hydrophilic oligo-isoprenoids results in modifications of the rubber structure and the synthesis of new material. In order to find an alternative process to recover the degradation products, a continuous extraction method using a biphasic system is described. The enzymatic activity of Lcp1VH2 was studied in the presence of ethyl acetate and pentane as extraction solvents. Oligo(cis-1,4-isoprene) molecular species were isolated from the organic phase and analyzed by Electrospray Ionization Mass Spectrometry. The enzymatic reaction process was evaluated in terms of the biotransformation yield of poly(cis-1,4-isoprene) rubber into the corresponding degradation products. Biotransformation yields of between 42-52 % were achieved depending on the enzymatic reactor design and the extraction solvent. The results also showed that the mass distribution of the oligo(cis-1,4-isoprene) depended on the organic solvent applied. A novel, simple and effective process is demonstrated for biotransformation of poly(cis-1,4-isoprene) rubber with high oligo-isoprenoid molecules recovery yields.

Accumulation of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) by Azotobacter vinelandii with different 3HV fraction in shake flasks and bioreactor.

In the present study, the production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) by Azotobacter vinelandii was evaluated in shake flasks and bioreactors, utilizing different precursors and oxygen transfer rates (OTRs). In shake flask cultures, the highest PHBV yield from sucrose (0.16 g g-1) and 3-hydroxyvalerate (3HV) fraction in the PHA chain (27.4 mol%) were obtained with valerate (1.0 g L-1). In the bioreactor, the cultures were grown under oxygen-limited conditions, and the maximum OTR (OTRmax) was varied by adjusting the agitation rate. In the cultures grown at low OTRmax (4.3 mmol L-1 h-1), the intracellular content of PHBV (73% w w-1) was improved, whereas a maximum 3HV fraction (35 mol %) was obtained when a higher OTRmax (17.2 mmol L-1 h-1, to 600 rpm) was employed. The findings obtained suggest that the PHBV production and the content of 3HV incorporated into the polymer were affected by the OTR. Based on the evidence, it is possible to produce PHBV with a different composition by varying the OTR of the culture; thus, the approach in this study could be used to scale up PHBV production.

Coenzyme Q production by metabolic engineered Escherichia coli strains in defined medium.

Coenzyme Q (CoQ) plays an important role as an electron transporter in the respiratory chain. It is formed from a benzoquinone ring and an isoprenoid chain of a specific length depending on the organism. We constructed an engineered Escherichia coli strain (menF) unable to produce demethylmenaquinone and menaquinone, compounds that compete for both chorismate, precursor of the benzoquinone ring, and the isoprenoid chain involved in CoQ biosynthesis. In addition, a mutant strain (entC) unable to produce enterobactin, high-affinity siderophore, synthesized from chorismate, and a double mutant (entC-menF) were constructed. The use of glucose or glycerol as carbon sources was also evaluated for the production of CoQ8 in these strains. The double mutant (entC-menF) showed 18% increase in CoQ8-specific content compared to the control strain; however, the single-mutant strains did not show statistically significant differences in CoQ8-specific content respect to the control, in glucose medium in bioreactor experiments. Glycerol was significantly superior compared to glucose for the production of CoQ8 in E. coli, where the CoQ8-specific content increased 126% and 53% in the control and double-mutant strain, respectively. The expression of genes related to CoQ8 biosynthesis is reported, where the entC-menF double-mutant strain showed a significant increase in the expression of CoQ8 biosynthesis-related genes when glycerol was used as sole carbon source. The control strain did not show gene expression difference between both carbon sources, indicating a possible regulation at a different level.

Poly(3-hydroxybutyrate) accumulation by Azotobacter vinelandii under different oxygen transfer strategies.

Azotobacter vinelandii OP is a bacterium that produces poly(3-hydroxybutyrate) (PHB). PHB production in a stirred bioreactor, at different oxygen transfer strategies, was evaluated. By applying different oxygen contents in the inlet gas, the oxygen transfer rate (OTR) was changed under a constant agitation rate. Batch cultures were performed without dissolved oxygen tension (DOT) control (using 9% and 21% oxygen in the inlet gas) and under DOT control (4%) using gas blending. The cultures that developed without DOT control were limited by oxygen. As result of varying the oxygen content in the inlet gas, a lower OTR (4.6 mmol L-1 h-1) and specific oxygen uptake rate (11.6 mmol g-1 h-1) were obtained using 9% oxygen in the inlet gas. The use of 9% oxygen in the inlet gas was the most suitable for improving the intracellular PHB content (56 ± 6 w w-1). For the first time, PHB accumulation in A. vinelandii OP cultures, developed with different OTRs, was compared under homogeneous mixing conditions, demonstrating that bacterial respiration affects PHB synthesis. These results can be used to design new oxygen transfer strategies to produce PHB under productive conditions.

Bacterial alginate production: an overview of its biosynthesis and potential industrial production.

Alginate is a linear polysaccharide that can be used for different applications in the food and pharmaceutical industries. These polysaccharides have a chemical structure composed of subunits of (1-4)-ß-D-mannuronic acid (M) and its C-5 epimer α-L-guluronic acid (G). The monomer composition and molecular weight of alginates are known to have effects on their properties. Currently, these polysaccharides are commercially extracted from seaweed but can also be produced by Azotobacter vinelandii and Pseudomonas spp. as an extracellular polymer. One strategy to produce alginates with different molecular weights and with reproducible physicochemical characteristics is through the manipulation of the culture conditions during fermentation. This mini-review provides a comparative analysis of the metabolic pathways and molecular mechanisms involved in alginate polymerization from A. vinelandii and Pseudomonas spp. Different fermentation strategies used to produce alginates at a bioreactor laboratory scale are described.

Molecular mass of Poly-3-hydroxybutyrate (P3HB) produced by Azotobacter vinelandii is influenced by the polymer content in the inoculum.

Poly-3-hydroxybutyrate (P3HB) is a biopolymer produced by Azotobacter vinelandii. The physicochemical properties and applications of P3HB are strongly influenced by its weight-average molecular mass (Mw), and in A. vinelandii, it could be influenced by the culture conditions. The aim of this study was to evaluate the effect of the P3HB content of the inoculum on the Mw of the polymer produced by A. vinelandii OP in bioreactor cultures. A. vinelandii cells containing 20, 50 and 70% of P3HB were used as inoculum. The P3HB content in the inoculum affected the volumetric P3HB productivity (qP3HB) and the Mw of P3HB. Those cultures inoculated with cells containing 20% of P3HB, achieved the highest qP3HB (0.17±0.018gP3HBL-1h-1); whereas a P3HB content of 70% was reflected as a low qP3HB (0.021±0.002gP3HBL-1h-1). On the other hand, using an inoculum with 70% of polymer content, the Mw of the biopolymer remained stable at values close to 3200kDa; whereas, when an inoculum with 20% of P3HB was used, the Mw decreased drastically during early stages of cultivation. These results show that manipulating the P3HB content of the inoculum is possible to produce biopolymers with a suitable Mw.

Different responses in the expression of alginases, alginate polymerase and acetylation genes during alginate production by Azotobacter vinelandii under oxygen-controlled conditions.

Alginate production and gene expression of genes involved in alginate biosynthesis were evaluated in continuous cultures under dissolved oxygen tension (DOT) controlled conditions. Chemostat at 8% DOT showed an increase in the specific oxygen uptake rate [Formula: see text] from 10.9 to 45.3 mmol g-1 h-1 by changes in the dilution rate (D) from 0.06 to 0.10 h-1, whereas under 1% DOT the [Formula: see text] was not affected. Alginate molecular weight was not affected by DOT. However, chemostat at 1% DOT showed a downregulation up to 20-fold in genes encoding both the alginate polymerase (alg8, alg44), alginate acetylases (algV, algI) and alginate lyase AlgL. alyA1 and algE7 lyases gene expressions presented an opposite behavior by changing the DOT, suggesting that A. vinelandii can use specific depolymerases depending on the oxygen level. Overall, the DOT level have a differential effect on genes involved in alginate synthesis, thus a gene expression equilibrium determines the production of alginates of similar molecular weight under DOT controlled.

A comparative study of glycerol and sorbitol as co-substrates in methanol-induced cultures of Pichia pastoris: temperature effect and scale-up simulation.

The production of recombinant proteins by Pichia pastoris under AOX1 promoter is usually performed using methanol together with either glycerol or sorbitol as co-substrate. Although both co-substrates have been widely used, comparative studies are scarce. In addition, these comparisons have been performed at different specific growth rate (µ) that it is well known that has an important effect on productivity. Thus, the effect of using these co-substrates on the production of Rhyzopus oryzae lipase (ROL) by P. pastoris was compared in continuous cultures growing at the same µ at either 22 or 30 °C. Results show that using glycerol as co-substrate led to higher volumetric productivities, and lower specific and volumetric methanol consumption rates. Scale-up simulation with 10-10,000 L bioreactor sizes indicated that glycerol produced the highest volumetric productivity of ROL with lower aeration requirements. Therefore, glycerol rises as a better option than sorbitol in ROL production.