Stimulation of reductive glycerol metabolism by overexpression of an aldehyde dehydrogenase in a recombinant <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis> strain defective in the oxidative pathway

Previously, we constructed a glycerol oxidative pathway-deficient mutant strain of Klebsiella pneumoniae by inactivation of glycerol dehydrogenase (dhaD) to eliminate by-product synthesis during production of 1,3-propanediol (1,3-PD) from glycerol. Although by-product formation was successfully blocked in the resultant strain, the yield of 1,3-PD was not enhanced, probably because dhaD disruption resulted in insufficient regeneration of the cofactor NADH essential for the activity of 1,3-PD oxidoreductase (DhaT). To improve cofactor regeneration, in the present study we overexpressed an NAD⁺-dependent aldehyde dehydrogenase in the recombinant strain. To this end, an aldehyde dehydrogenase AldHk homologous to E. coli AldH but with NAD⁺-dependent propionaldehyde dehydrogenase activity was identified in K. pneumoniae. Functional analysis revealed that the substrate specificity of AldHk embraced various aldehydes including propionaldehyde, and that NAD⁺ was preferred over NADP⁺ as a cofactor. Overexpression of AldHk in the glycerol oxidative pathway-deficient mutant AK/pVOTHk resulted in a 3.6-fold increase (0.57 g l⁻¹ to 2.07 g l⁻¹) in the production of 3-hydroxypropionic acid (3-HP), and a 1.1-fold enhancement (8.43 g l⁻¹ to 9.65 g l⁻¹) of 1,3-PD synthesis, when glycerol was provided as the carbon source, compared to the levels synthesized by the control strain (AK/pVOT). Batch fermentation using AK/pVOTHk showed a significant increase (to 70%, w/w) in conversion of glycerol to the reductive metabolites, 1,3-PD and 3-HP, with no production of by-products except acetate.     

Production of 3-hydroxypropionic acid through propionaldehyde dehydrogenase PduP mediated biosynthetic pathway in Klebsiella pneumoniae

The pduP gene encodes a propionaldehyde dehydrogenase (PduP) was investigated for the role in 3-hydroxypropionic acid (3-HP) glycerol metabolism in Klebsiella pneumoniae. The enzyme assay showed that cell extracts from a pduP mutant strain lacked measurable dehydrogenase activity. Additionally, the mutant strain accumulated the cytotoxic intermediate metabolite 3-hydroxypropionaldehyde (3-HPA), causing both cell death and a lower final 3-HP titer. Ectopic expression of pduP restored normal cell growth to mutant. The enzymatic property of recombinant protein from Escherichia coli was examined, exhibiting a broad substrate specificity, being active on 3-HPA. The present work is thus the first to demonstrate the role of PduP in glycerol metabolism and biosynthesis of 3-HP.     

PduP is a coenzyme-a-acylating propionaldehyde dehydrogenase associated with the polyhedral bodies involved in B<Subscript>12</Subscript>-dependent 1,2-propanediol degradation by<Emphasis Type="Italic"> Salmonella enterica</Emphasis> serovar Typhimurium LT2

Salmonella enterica forms polyhedral bodies involved in coenzyme-B₁₂-dependent 1,2-propanediol degradation. Prior studies showed that these bodies consist of a proteinaceous shell partly composed of the PduA protein, coenzyme-B₁₂-dependent diol dehydratase, and additional unidentified proteins. In this report, we show that the PduP protein is a polyhedral-body-associated CoA-acylating aldehyde dehydrogenase important for 1,2-propanediol degradation by S. enterica. A PCR-based method was used to construct a precise nonpolar deletion of the gene pduP. The resulting pduP deletion strain grew poorly on 1,2-propanediol minimal medium and expressed 105-fold less propionaldehyde dehydrogenase activity (0.011 mol min^–1 mg^–1) than did wild-type S. enterica grown under similar conditions (1.15 mol min^–1 mg^–1). An Escherichia coli strain was constructed for high-level production of His₈-PduP, which was purified by nickel-affinity chromatography and shown to have 15.2 mol min^–1 mg^–1 propionaldehyde dehydrogenase activity. Analysis of assay mixtures by reverse-phase HPLC and mass spectrometry established that propionyl-CoA was the product of the PduP reaction. For subcellular localization, purified His₈-PduP was used as antigen for the preparation of polyclonal antiserum. The antiserum obtained was shown to have high specificity for the PduP protein and was used in immunogold electron microscopy studies, which indicated that PduP was associated with the polyhedral bodies involved in 1,2-propanediol degradation. Further evidence for the localization of the PduP enzyme was obtained by showing that propionaldehyde dehydrogenase activity co-purified with the polyhedral bodies. The fact that both Ado-B₁₂-dependent diol dehydratase and propionaldehyde dehydrogenase are associated with the polyhedral bodies is consistent with the proposal that these structures function to minimize propionaldehyde toxicity during the growth of S. enterica on 1,2-propanediol.     

High alcohol production by repeated batch fermentation using an immobilized osmotolerant Saccharomyces cerevisiae

A repeated batch fermentation system was used to produce ethanol using an osmotolerant Saccharomyces cerevisiae (VS₃) immobilized in calcium alginate beads. For comparison free cells were also used to produce ethanol by repeated batch fermentation. Fermentation was carried for six cycles with 125, 250 or 500 beads using 150, 200 or 250 g glucose L⁻¹ at 30°C. The maximum amount of ethanol produced by immobilized VS₃ using 150 g L⁻¹ glucose was only 44 g L⁻¹ after 48 h, while the amount of ethanol produced by free cells in the first cycle was 72 g L⁻¹. However in subsequent fed batch cultures more ethanol was produced by immobilized cells compared to free cells. The amount of ethanol produced by free cells decreased from 72 g L⁻¹ to 25 g L⁻¹ after the fourth cycle, while that of immobilized cells increased from 44 to 72 g L⁻¹. The maximum amount of ethanol produced by immobilized VS₃ cells using 150, 200 and 250 g glucose L⁻¹ was 72.5, 93 and 87 g ethanol L⁻¹ at 30°C. Journal of Industrial Microbiology & Biotechnology (2000) 24, 222–226. Received 16 September 1999/ Accepted in revised form 22 December 1999     

Production of ��-poly-l-lysine using a novel two-stage pH control strategy by Streptomyces sp. M-Z18 from glycerol

The production of ε-poly-l-lysine (ε-PL) by Streptomyces sp. M-Z18 from glycerol was investigated in a 5-L jar-fermenter. Batch fermentations by Streptomyces sp. M-Z18 at various pH values ranging from 3.5 to 4.5 were studied. Based on the analysis of the time course of specific cell growth rate and specific ε-PL formation rate, a novel two-stage pH control strategy was developed to improve ε-PL production by shifting the culture pH from 3.5 to 3.8 after 36 h of cultivation. By applying the strategy, the maximal ε-PL concentration and productivity had a significant improvement and reached 9.13 g L⁻¹ and 4.76 g L⁻¹ day⁻¹, respectively, compared with those in one-stage pH control process where the pH value is controlled at 3.5 (7.83 g L⁻¹ and 3.13 g L⁻¹ day⁻¹). Fed-batch fermentation with two-stage pH control strategy was also applied to produce ε-PL; final ε-PL concentration of 30.11 g L⁻¹ was obtained, being 3.3-fold greater than that of batch fermentation. To our knowledge, it is the first report on production of ε-PL from glycerol in fermenter scale and achievement of high ε-PL production with two-stage pH control strategy.     

Production of d-arabitol by a newly isolated Kodamaea ohmeri

A new yeast, isolated from natural osmophilic sources, produces d-arabitol as the main metabolic product from glucose. According to 18S rRNA analysis, the NH-9 strain belongs to the genus Kodamaea. The optimal culture conditions for inducing production of d-arabitol were 37 °C, neutral pH, 220 rpm shaking, and 5% inoculum. The yeast produced 81.2 ± 0.67 g L⁻¹ d-arabitol from 200 g L⁻¹ d-glucose in 72 h with a yield of 0.406 g g⁻¹ glucose and volumetric productivity Q_\textP Q_{\text{P}} of 1.128 g L⁻¹ h⁻¹. Semi-continuous repeated-batch fermentation was performed in shaker-flasks to enhance the process of d-arabitol production by Kodamaea ohmeri NH-9 from d-glucose. Under repeated-batch culture conditions, the highest volumetric productivity was 1.380 g L⁻¹ h⁻¹.     

Production of Cold-Adapted Amylase by Marine Bacterium <Emphasis Type="Italic">Wangia</Emphasis> sp. C52: Optimization,Modeling, and Partial Characterization

The aim of this work was to optimize the fermentation parameters in the shake-flask culture of marine bacterium Wangia sp. C52 to increase cold-adapted amylase production using two statistical experimental methods including Plackett–Burman design, which was applied to find the key ingredients for the best medium composition, and response surface methodology, which was used to determine the optimal concentrations of these components. The results showed starch, tryptone, and initial pH had significant effects on the cold-adapted amylase production. A central composite design was then employed to further optimize these three factors. The experimental results indicated that the optimized composition of medium was 6.38 g L⁻¹ starch, 33.84 g L⁻¹ tryptone, 3.00 g L⁻¹ yeast extract, 30 g L⁻¹ NaCl, 0.60 g L⁻¹ MgSO₄ and 0.56 g L⁻¹ CaCl₂. The optimized cultivation conditions for amylase production were pH 7.18, a temperature of 20°C, and a shaking speed of 180 rpm. Under the proposed optimized conditions, the amylase experimental yield (676.63 U mL⁻¹) closely matched the yield (685.60 U mL⁻¹) predicted by the statistical model. The optimization of the medium contributed to tenfold higher amylase production than that of the control in shake-flask experiments.     

Production of hexanoic acid from d-galactitol by a newly isolated Clostridium sp. BS-1

In a study screening anaerobic microbes utilizing d-galactitol as a fermentable carbon source, four bacterial strains were isolated from an enrichment culture producing H₂, ethanol, butanol, acetic acid, butyric acid, and hexanoic acid. Among these isolates, strain BS-1 produced hexanoic acid as a major metabolic product of anaerobic fermentation with d-galactitol. Strain BS-1 belonged to the genus Clostridium based on phylogenetic analysis using 16S rRNA gene sequences, and the most closely related strain was Clostridium sporosphaeroides DSM 1294^T, with 94.4% 16S rRNA gene similarity. In batch cultures, Clostridium sp. BS-1 produced 550 ± 31 mL L⁻¹ of H₂, 0.36 ± 0.01 g L⁻¹ of acetic acid, 0.44 ± 0.01 g L⁻¹ of butyric acid, and 0.98 ± 0.03 g L⁻¹ of hexanoic acid in a 4-day cultivation. The production of hexanoic acid increased to 1.22 and 1.73 g L⁻¹ with the addition of 1.5 g L⁻¹ of sodium acetate and 100 mM 2-(N-morpholino)ethanesulfonic acid (MES), respectively. Especially when 1.5 g L⁻¹ of sodium acetate and 100 mM MES were added simultaneously, the production of hexanoic acid increased up to 2.99 g L⁻¹. Without adding sodium acetate, 2.75 g L⁻¹ of hexanoic acid production from d-galactitol was achieved using a coculture of Clostridium sp. BS-1 and one of the isolates, Clostridium sp. BS-7, in the presence of 100 mM MES. In addition, volatile fatty acid (VFA) production by Clostridium sp. BS-1 from d-galactitol and d-glucose was enhanced when a more reduced culture redox potential (CRP) was applied via addition of Na₂S·9H₂O.     

Cultivation of <Emphasis Type="Italic">Chlorella emersonii</Emphasis> with flue gas derived from a cement plant

The present study reviews the options of cultivating the green alga, Chlorella emersonii, under photoautotrophic conditions with flue gas derived from a cement plant. It was conducted in the Lafarge Perlmooser plant in Retznei, Austria, where stone coal and various surrogate fuels such as used tyres, plastics and meat-and-bone meal are incinerated for heating limestone. During 30 days of cultivation, flue gas had no visible adverse effects compared to the controls grown with pure CO₂. The semi-continuous cultivation with media recycling was performed in 5.5-L pH-stat photobioreactors. The essay using CO₂ from flue gas yielded a total of 2.00 g L⁻¹ microalgal dry mass and a CO₂ fixation of 3.25 g L⁻¹. In the control, a total of 2.06 g L⁻¹ dry mass was produced and 3.38 g L⁻¹ CO₂ was fixed. Mean growth rates were between 0.10 day⁻¹ (control) and 0.13 day⁻¹ (flue gas). No accumulation of flue gas residues was detected in the culture medium. At the end of the experiment, however, the concentration of lead was three times higher in algal biomass compared to the control, indicating that cultures aerated with this type of flue gas should not be used as food supplements or animal feed.     

Production of tetramethylpyrazine by batch culture of <Emphasis Type="Italic">Bacillus subtilis</Emphasis> with optimal pH control strategy

The effects of initial culture pH ranging from 5.0 to 7.5 on biomass content, precursor 3-hydroxy-2-butanone (HB) accumulation, and 2,3,5,6-tetramethylpyrazine (TTMP) formation by Bacillus subtilis CCTCC M 208157 were investigated in shake flask fermentation. Weak acidic conditions were found to favor cell growth and precursor HB accumulation, while TTMP could be synthesized more efficiently in conditions with initial pH towards neutrality. Batch bioprocess of TTMP fermentation by Bacillus subtilis CCTCC M 208157 at various controlled pH values ranging from 5.5 to 7.0 was then examined in 7.5-l fermentor. The results suggested that optimum pH for cell growth and precursor HB accumulation was 5.5 with maximum cell growth rate (Q _x) and precursor HB accumulation rate (Q _HB) of 0.833 g l⁻¹ h⁻¹ and 1.118 g l⁻¹ h⁻¹, respectively, while optimum pH for TTMP formation was 7.0 with maximum TTMP formation rate (Q _TTMP) of 0.095 g l⁻¹ h⁻¹. A pH-shifted strategy was accordingly developed to improve TTMP production in bioreactor fermentation by shifting the culture pH from 5.5 to 7.0 after 48 h of cultivation. By applying the strategy, final TTMP concentration of 7.43 g l⁻¹ was obtained, being 22.2% greater than that of constant-pH fermentation.     

Development of a cost-effective glucose-corn steep medium for production of butanol by Clostridium beijerinckii

Corn steep water (CSW) medium (1.6% solids plus 6% glucose) was evaluated for growth and butanol production by Clostridium beijerinckii NCIMB 8052 wild-type and hyper-amylolytic, hyper-butanol-producing mutant strain BA101. CSW alone was not a suitable substrate, whereas addition of glucose supported growth and butanol production by both strains. In a batch-scale fermentation using an optimized 6% glucose-1.6% solids CSW medium, C. beijerinckii NCIMB 8052 and strain BA101 produced 10.7 g L⁻¹ and 14.5 g L⁻¹ of butanol, respectively. The total solvents (acetone, butanol, and ethanol) produced by C. beijerinckii NCIMB 8052 and strain BA101 were 14 g L⁻¹ and 20 g L⁻¹, respectively. Initial fermentation in small-scale flasks containing 6% maltodextrin-1.6% solids concentration CSW medium resulted in 6 g L⁻¹ and 12.6 g L⁻¹ of butanol production by C. beijerinckii NCIMB 8052 and strain BA101, respectively. CSW can serve as an economic source of nitrogen, vitamins, amino acids, minerals, and other nutrients. Thus, it is feasible to use 6% glucose-1.6% solids CSW medium in place of semi-defined P2 medium. Received 9 February 1998/ Accepted in revised form 1 September 1998     

Simultaneous production and characterization of medium-chain-length polyhydroxyalkanoates and alginate oligosaccharides by <Emphasis Type="Italic">Pseudomonas mendocina</Emphasis> NK-01

When Pseudomonas mendocina NK-01 was cultivated in a 200-L fermentor using glucose as carbon source, 0.316 g L⁻¹ medium-chain-length polyhydroxyalkanoate (PHA_MCL) and 0.57 g L⁻¹ alginate oligosaccharides (AO) were obtained at the end of the process. GC/MS was used to characterize the PHA_MCL, which was found to be a polymer mainly consisting of 3HO (3-hydroxyoctanoate) and 3HD (3-hydroxydecanoate). T _m and T _g values for the PHA_MCL were 51.03°C and −41.21°C, respectively, by DSC. Its decomposition temperature was about 300°C. The elongation at break was 700% under 12 MPa stress. MS and GPC were also carried out to characterize the AO which had weight-average molecular weights of 1,546 and 1,029 Da, respectively, for the two main components at the end of the fermentation process. MS analysis revealed that the AO were consisted of β-d-mannuronic acid and/or α-l-guluronic acid, and the β-d-mannuronic acid and/or α-l-guluronic acid residues were partially acetylated at position C2 or C3.     

Continuous production of isopropanol and butanol using <Emphasis Type="Italic">Clostridium beijerinckii</Emphasis> DSM 6423

Clostridium beijerinckii DSM 6423 was studied using different continuous production methods to give maximum and stable production of isopropanol and n-butanol. In a single-stage continuous culture, when wood pulp was added as a cell holding material, we could increase the solvent productivity from 0.47 to 5.52 g L⁻¹ h⁻¹ with the yield of 54% from glucose. The overall solvent concentration of 7.51 g L⁻¹ (39.4% isopropanol and 60.6% n-butanol) with the maximum solvent productivity of 0.84 g L⁻¹ h⁻¹ was obtained with two-stage continuous culture. We were able to run the process for more than 48 overall retention times without losing the ability to produce solvents.     

Production of ectoine through a combined process that uses both growing and resting cells of <Emphasis Type="Italic">Halomonas salina</Emphasis> DSM 5928<Superscript>T</Superscript>

Using ectoine-excreting strain Halomonas salina DSM 5928^T, we developed a new process for high-efficiency production of ectoine, which involved a combined process of batch fermentation by growing cells and production by resting cells. In the first stage, batch fermentation was carried out using growing cells under optimal fermentation conditions. The second stage was the production phase, in which ectoine was synthesized and excreted by phosphate-limited resting cells. Optimal conditions for synthesis and excretion of ectoine during batch fermentation in a 10 l fermentor were 0.5 mol l⁻¹ NaCl and an initial monosodium glutamate concentration of 80 g l⁻¹ respectively. The pH was adjusted to 7.0 and the temperature was maintained at 33°C. In phosphate-limited resting cells medium, monosodium glutamate and NaCl concentration was 200 g l⁻¹ and 0.5 mol l⁻¹, respectively, as well as pH was 7.0. The total concentration of ectoine produced was 14.86 g l⁻¹, the productivity and yield of ectoine was 7.75 g l⁻¹ day⁻¹ and 0.14 g g⁻¹, respectively, and the percentage of ectoine excreted was 79%. These levels of ectoine production and excretion are the highest reported to date.     

Somatic embryogenesis and plant regeneration in oil palm using the thin cell layer technique

An efficient procedure has been developed for inducing somatic embryogenesis and regeneration of plants from tissue cultures of oil palm (Elaeis guineensis Jacq.). Thin transverse sections (thin cell layer explants) of different position in the shoot apex and leaf sheath of oil palm were cultivated in Murashige and Skoog (MS) (Physiol Plant 15:473–497, 1962) medium supplemented with 0–450 μM picloram and 2,4-D with 3.0% sucrose, 500 mg L⁻¹ glutamine, and 0.3 g L⁻¹ activated charcoal and gelled with 2.5 g L⁻¹ Phytagel. Embryogenic calluses were evaluated 12 wk after inoculation. Picloram (450 μM) was effective in inducing embryogenic calluses in 41.5% of the basal explants. Embryogenic calluses were maintained on a maturation medium composed of basal media, plus 0.6 μM NAA and 12.30 μM 2iP, 0.3 g L⁻¹ activated charcoal, and 500 mg L⁻¹ glutamine, with subcultures at 4-wk intervals. Somatic embryos were converted to plants on MS medium with macro- and micronutrients at half-strength, 2% sucrose, and 1.0 g L⁻¹ activated charcoal and gelled with 2.5 g L⁻¹ Phytagel.     

Continuous butanol production with reduced byproducts formation from glycerol by a hyper producing mutant of <Emphasis Type="Italic">Clostridium pasteurianum</Emphasis>

Butanol, a four-carbon primary alcohol (C₄H₁₀O), is an important industrial chemical and has a good potential to be used as a superior biofuel. Bio-based production of butanol from renewable feedstock is a promising and sustainable alternative to substitute petroleum-based fuels. Here, we report the development of a process for butanol production from glycerol, which is abundantly available as a byproduct of biodiesel production. First, a hyper butanol producing strain of Clostridium pasteurianum was isolated by chemical mutagenesis. The best mutant strain, C. pasteurianum MBEL_GLY2, was able to produce 10.8 g l⁻¹ butanol from 80 g l⁻¹ glycerol as compared to 7.6 g l⁻¹ butanol produced by the parent strain. Next, the process parameters were optimized to maximize butanol production from glycerol. Under the optimized batch condition, the butanol concentration, yield, and productivity of 17.8 g l⁻¹, 0.30 g g⁻¹, and 0.43 g l⁻¹ h⁻¹ could be achieved. Finally, continuous fermentation of C. pasteurianum MBEL_GLY2 with cell recycling was carried out using glycerol as a major carbon source at several different dilution rates. The continuous fermentation was run for 710 h without strain degeneration. The acetone–butanol–ethanol productivity and the butanol productivity of 8.3 and 7.8 g l⁻¹ h⁻¹, respectively, could be achieved at the dilution rate of 0.9 h⁻¹. This study reports continuous production of butanol with reduced byproducts formation from glycerol using C. pasteurianum, and thus could help design a bioprocess for the improved production of butanol.     

Application of oscillation for efficiency improvement of continuous ethanol fermentation with Saccharomyces cerevisiae under very-high-gravity conditions

Compared with steady state, oscillation in continuous very-high-gravity ethanol fermentation with Saccharomyces cerevisiae improved process productivity, which was thus introduced for the fermentation system composed of a tank fermentor followed by four-stage packed tubular bioreactors. When the very-high-gravity medium containing 280 g l⁻¹ glucose was fed at the dilution rate of 0.04 h⁻¹, the average ethanol of 15.8% (v/v) and residual glucose of 1.5 g l⁻¹ were achieved under the oscillatory state, with an average ethanol productivity of 2.14 g h⁻¹ l⁻¹. By contrast, only 14.8% (v/v) ethanol was achieved under the steady state at the same dilution rate, and the residual glucose was as high as 17.1 g l⁻¹, with an ethanol productivity of 2.00 g h⁻¹ l⁻¹, indicating a 7% improvement under the oscillatory state. When the fermentation system was operated under the steady state at the dilution rate of 0.027 h⁻¹ to extend the average fermentation time to 88 h from 59 h, the ethanol concentration increased slightly to 15.4% (v/v) and residual glucose decreased to 7.3 g l⁻¹, correspondingly, but the ethanol productivity was decreased drastically to 1.43 g h⁻¹ l⁻¹, indicating a 48% improvement under the oscillatory state at the dilution rate of 0.04 h⁻¹.     

Environmental conditions during glycerol bioconversion affect 3-hydroxypropionic acid bioproduction by Limosilactobacillus reuteri DSM 17938

3-Hydroxypropionic acid (3-HP) is a platform molecule whose biological production was carried out by the bacterium Limosilactobacillus reuteri according to a two-step process: first, a growth phase in batch mode on glucose, then a glycerol bioconversion into 3-HP in fed-batch mode. With the objective of improving 3-HP bioproduction, this study aimed at defining the operating conditions during the bioconversion phase that increases the bioproduction performance. A central composite rotatable design allowed testing various pH levels and specific glycerol feeding rates. By establishing response surfaces, optimal conditions have been identified that were different depending on the considered output variable (final 3-HP quantity, 3-HP production yield and production rate). Of them, 3-HP final quantity and 3-HP production yield were maximized at pH 6.0 and at specific glycerol feeding rates of 60 and 55 mg_gly g_CDW⁻¹ h⁻¹, respectively. The specific 3-HP production rate was the highest at the upper limit of the specific substrate feeding rate (80 mg_gly g_CDW⁻¹ h⁻¹) but was not affected by the pH. An additional experiment was carried out at pH 6.0 and a specific glycerol feeding rate of 80 mg_gly g_CDW⁻¹ h⁻¹ to validate the previous observations. In conclusion, the results showed a significant improvement of 3-HP concentration by 13%, of specific production rate by 34% and of 3-HP volumetric productivity by 39%, as compared to the initial values.     

1,3-Propandiol production by engineered Hansenula polymorpha expressing dha genes from Klebsiella pneumoniae

Currently, 1,3-propanediol (1,3-PD) is an important chemical widely used in polymer production, but its availability is being restricted owing to its expensive chemical synthesis. A methylotrophic yeast Hansenula polymorpha was engineered by expression of dhaB1, dhaB2, dhaB3, dhaB _RA1 and dhaB _RA2 encoding glycerol dehydratase complex and dhaT encoding 1,3-PD oxidoreductase from Klebsiella pneumoniae under direction of promoter of glyceraldehyde-3 phosphate dehydrogenase (GAPDH). The engineered recombinant yeast strain can produce 1,3-PD from glucose (2.4 g L⁻¹) as well as glycerol (0.8 g L⁻¹), which might lead to a safe and cost-effective method for industrial production of 1,3-PD from various biomass resources.     

Sucrose regulated enhanced induction of anthraquinone,phenolics, flavonoids biosynthesis and activities of antioxidant enzymes in adventitious root suspension cultures of <Emphasis Type="Italic">Morinda citrifolia</Emphasis> (L.)

The effect of initial sucrose concentration was investigated in root suspension cultures of Morinda citrifolia to improve root growth and secondary metabolites production, i.e. anthraquinone, phenolics and flavonoids. Besides, oxidative stress level, antioxidant enzymes activity and membranes damage under different sucrose concentration were estimated. A 5% sucrose supply was shown to be optimal for the production of root dry mass, but higher sucrose concentrations of 7–9% inhibited the accumulation of root dry weight (DW). However, the maximum production of anthraquinone (251.89 g L⁻¹ DW), phenolics (165.14 g L⁻¹ DW) and flavonoids (163.56 g L⁻¹ DW) were achieved at 1% sucrose-treated culture, which may be a source carbon skeletons for secondary metabolism. At the same time was observed low oxidative damage, which could be associated with high levels of secondary metabolites and the increased activity of catalase. Although, catalase (CAT) activity were stimulated at 7–9% sucrose-treated cultures, high accumulation of hydrogen peroxide (H₂O₂) and peroxidation of lipid (MDA) was induced. The observed high activity of CAT and guaiacol peroxidase (G-POD) were not sufficient enough to mitigate the toxic effect of H₂O₂.