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Gordonia alkanivorans strain 1B is a desulfurizing bacterium and a hyper-pigment producer. Most carotenoid optimization studies have been performed with light, but little is still known on how carbon/sulfur-source concentrations influence carotenoid production under darkness. In this work, a surface response methodology based on a two-factor Doehlert distribution (% glucose in a glucose/fructose 10 g/L mixture; sulfate concentration) was used to study carotenoid and biomass production without light. These responses were then compared to those previously obtained under light. Moreover, carbon consumption was also monitored, and different metabolic parameters were further calculated. The results indicate that both light and glucose promote slower growth rates, but stimulate carotenoid production and carbon conversion to carotenoids and biomass. Fructose induces higher growth rates, and greater biomass production at 72 h; however, its presence seems to inhibit carotenoid production. Moreover, although at a much lower yield than under light, results demonstrate that under darkness the highest carotenoid production can be achieved with 100% glucose (10 g/L), ≥27 mg/L sulfate, and high growth time (>216 h). These results give a novel insight into the metabolism of strain 1B, highlighting the importance of culture conditions optimization to increase the process efficiency for carotenoid and/or biomass production.

The optimization of an alkaline pretreatment process for the delignification of sugarcane bagasse (SCB) to enhance the subsequent enzymatic hydrolysis was performed according to the Doehlert uniform shell design. In this experimental design, the effect of two factors—potassium hydroxide (KOH) concentration and autoclaving time at 121 C (1 atm)—on cellulose, hemicellulose, or the total polysaccharide and lignin content in SCB was evaluated. This response surface methodology revealed that KOH concentration is the factor that most influences the chemical characteristics of treated SCB (SCBt), with optimal conditions for the highest delignification being KOH in the range 5–10% (w/v) and an autoclaving time of 35 min, which provides an average of 97% total polysaccharides without inhibitor accumulation (furfural, 5-hydroxymethyl furfural) and #5% lignin. SCBt samples from two pretreatment conditions (KOH 3.25% – 13 min; KOH 10% – 35 min) were selected, based on the greatest delignification (70–74%) and polysaccharide availability (95–97%) after pretreatment, and further hydrolysed for fermentable sugar production. High sugar yields were obtained from both the pretreated samples (866 to 880 mg sugar per g biomass, respectively) in contrast with the 129 mg sugar per g raw biomass obtained from untreated SCB. These results demonstrate the effectiveness of KOH alkali pretreatments, which improves the overall digestibility of raw SCB polysaccharides from about 18% up to 91%. However, harsh alkali treatment (KOH 10%) is the most effective if the highest glucose/xylose ratio in the final sugar-rich hydrolysate is the aim. Hence, the use of sugar-rich hydrolysates obtained from SCBt as the carbon source for industrial purposes may provide a sustainable and economic solution for the production of bio-based added-value products, such as second generation (2G) bioethanol.

Aims Through biodesulphurization (BDS) is possible to remove the sulphur present in fossil fuels to carry out the very strict legislation. However, this biological process is limited by the cost of the culture medium, and thus, it is important to explore cheaper alternative carbon sources, such as Jerusalem artichoke (JA). These carbon sources usually contain sulphates which interfere with the BDS process. The goal of this work was to remove the sulphates from Jerusalem artichoke juice (JAJ) through BaCl2 precipitation viewing the optimization of dibenzothiophene (DBT) desulphurization by Gordonia alkanivorans strain 1B. Methods and Results Using a statistical design (Doehlert distribution), the effect of BaCl2 concentration (0·125–0·625%) and pH (5–9) was studied on sulphate concentration in hydrolysed JAJ. A validated surface response derived from data indicated that zero sulphates can be achieved with 0·5–0·55% (w/v) BaCl2 at pH 7; however, parallel BDS assays showed that the highest desulphurization was obtained with the juice treated with 0·5% (w/v) BaCl2 at pH 8·73. Further assays demonstrated that enhanced DBT desulphurization was achieved using hydrolysed JAJ treated in these optimal conditions. A total conversion of 400 μmol l−1 DBT into 2-hydroxybiphenyl (2-HBP) in

With the increasing awareness on the toxicity of several synthetic dyes, demand for pigments from natural sources, such as microbial carotenoids, has gained interest as a promising safe alternative colour additive. In this study, a surface response methodology based on the Doehlert distribution for two factors [% of glucose in a mixture of glucose + fructose (10 g/L total sugars), and sulfate concentration] was used towards the optimal carotenoids production by Gordonia alkanivorans strain 1B in the presence of light (400 lx). Time influence on pigment production by this bacterium was also evaluated, as well as the cell viability profile during longer incubation periods at optimal conditions. Indeed, the highest carotenoid production (2596-3100 mu g/g(DCW)) was obtained when strain 1B was cultivated in the optimal conditions: glucose 10 g/L and sulfate >= 22 mg/L, in the presence of light for 19 days at 30 degrees C, 150 rpm. Flow cytometry showed that the highest production was somehow related with the cellular stress. These results highlight the great potential of strain 1B as a new hyperpigment producer to be exploited towards several applications. info:eu-repo/semantics/publishedVersion

Biopesticides based on natural endophytic bacteria to control plant diseases are an ecological alternative to the chemical treatments. Bacillus species produce a wide variety of metabolites with biological activity like iturinic lipopeptides. This work addresses the production of biopesticides based on natural endophytic bacteria, isolated from Quercus suber. Artificial Neural Networks were used to maximize the percentage of inhibition triggered by antifungal activity of bioactive compounds produced by Bacillus amyloliquefaciens. The active compounds, produced in liquid cultures, inhibited the growth of fifteen fungi and exhibited a broader spectrum of antifungal activity against surface contaminant fungi, blue stain fungi and phytopathogenic fungi. A 19-7-6-1 neural network was selected to predict the percentage of inhibition produced by antifungal bioactive compounds. A good match among the observed and predicted values was obtained with the R2 values varying between 0.9965 – 0.9971 and 0.9974 – 0.9989 for training and test sets. The 19-7-6-1 neural network was used to establish the dilution rates that maximize the production of antifungal bioactive compounds, namely 0.25 h-1 for surface contaminant fungi, 0.45 h-1 for blue stain fungi and between 0.30 and 0.40 h-1 for phytopathogenic fungi. Artificial neural networks show great potential in the modelling and optimization of these bioprocesses., Les biopesticides à base de bactéries endophytes naturelles pour lutter contre les maladies des plantes constituent une alternative écologique aux traitements chimiques. Les espèces de Bacillus produisent une grande variété de métabolites biologiquement actifs tels que les lipopeptides ituriniques. Cette étude porte sur la production de biopesticides par des bactéries endophytes naturelles isolées du Quercus suber L. Des réseaux neuronaux artificiels ont été utilisés pour maximiser le pourcentage d’inhibition provoquée par l’activité antifongique des composés bioactifs produits par Bacillus amyloliquefaciens. Les composés actifs, produits en culture liquide, ont inhibé la croissance de 15 champignons et avaient un spectre d’activé antifongique plus large contre les contaminants fongiques de surface, les champignons de bleuissement et les champignons phytopathogènes. Un réseau neuronal 19-7-6-1 a été choisi pour prédire le pourcentage d’inhibition produit par les composés bioactifs antifongiques. Une bonne concordance entre les valeurs observées et prédites a été obtenue; les valeurs de R2 variaient de 0,9965 a` 0,9971 et de 0,9974 a` 0,9989 pour les bases d’apprentissage et de test. Le réseau neuronal 19-7-6-1 a été utilisé pour établir les taux de dilution qui maximisent la production des composés bioactifs antifongiques, nommément 0,25 h−1 pour les contaminants fongiques de surface, 0,45 h−1 pour les champignons de bleuissement et entre 0,30 et 0,40 h−1 pour les champignons phytopathogènes. Les réseaux neuronaux artificiels ont un potentiel élevé pour modéliser et optimiser ces processus biologiques.

Carob (Ceratonia siliqua L.) kibbles, a by-product of the carob bean gum production, were extracted by decoction and evaluated for its phenolic profile and potential use as an added-value antioxidant-rich product. The effect of decoction time (8–20 min) and temperature (80–100 °C) on the polyphenols content and overall antioxidant capacity expressed as percentage inhibition of the DPPH and ABTS free radicals, was performed using a Doehlert experimental design to assess the antioxidants extraction efficiency. It was observed that temperature had the most significant effect. The best conditions for total polyphenols and tannins extraction were 98.5 °C and 17 min, where ca. 39.5 mg GAE/g kibbles and 25.8 mg GAE/g kibbles on a dry basis were obtained, respectively. Under these conditions, inhibition of DPPH and ABTS radicals were 85% and 90%, respectively, showing the promising use of this biomass as an antioxidant source, which could be used, for example, in functional beverages.

Biodesulfurization can be a complementary technology to the hydrodesulfurization, the commonly physical-chemical process used for sulfur removal from crude oil. The desulfurizing bacterium Gordonia alkanivorans strain 1B as a fructophilic microorganism requires fructose as C-source. In this context, the main goal of this work was the optimization of a simultaneous saccharification and fermentation (SSF) approach using the Zygosaccharomyces bailii strain Talf1 crude enzymes with invertase activity and sucrose as a cheaper fructose-rich commercial C-source (50% fructose) towards dibenzothiophene (DBT) desulfurization by strain 1B. The determination of optimal conditions, for both sucrose hydrolysis and DBT desulfurization was carried out through two sequential experimental uniform designs according to the Doehlert distribution for two factors: pH (5.5-7.5) and temperature (28-38 °C), with the enzyme load of 1.16 U/g/L; and enzyme load (0-4 U/g/L) and temperature (28-38 °C), with pH at 7.5. Based on 2-hydroxybiphenyl production, the analysis of the response surfaces obtained pointed out for pH 7.5, 32 °C and 1.8 U/g/L as optimal conditions. Further optimized SSF of sucrose during the DBT desulfurization process permitted to attain a 4-fold enhanced biodesulfurization. This study opens a new focus of research through the exploitation of sustainable low cost sucrose-rich feedstocks towards a more economical viable bioprocess scale-up.

BACKGROUND: Biodesulfurization (BDS) is a complementary technology to hydrodesulfurization since it allows the removal of recalcitrant sulfur compounds present in fossil fuels. The cost of culture medium to produce the biocatalysts is still one limitation for BDS application. Carob pulp, as an alternative carbon source, can reduce this cost. However, the presence of sulfates is critical, since BDS is inhibited at very low concentrations. Thus, the goal of this work was to optimize the process of sulfur precipitation on carob pulp liquor. RESULT: The effect of BaCl2 concentration (0–0.5%) and exposure time (6–36 h) on sulfate removal from carob pulp liquor was studied according to a statistical design following the Doehlert distribution for two factors. This experimental design determined that 0.5% BaCl2 concentration for 21 h were adequate conditions for sulfate removal from carob pulp liquor using BDS. CONCLUSION: These results demonstrate that it is possible to use alternative carbon sources derived from agro-industrial wastes for BDS, even those with high sulfur levels. For future industrial application, an inexpensive culture medium would have to be employed in a large-scale process and carob pulp liquor could be the carbon source. © 2012 Society of Chemical Industry

Article history: Received 2 February 2010 Received in revised form 15 July 2010 Accepted 19 July 2010 Available online 27 July 2010 Keywords: Bacillus amiloliquefaciens Spore formation Anti-fungal activity Neural networks 1. Introduction Biopesticides based on natural endophytic bacteria to control plant diseases are a promising ecological alternative to chemical treatments. Bacillus species produce a wide variety of metabolites with interesting biological activities, among them iturinic lipopep- tides antibiotics (Bottone and Peluso, 2003; Cho et al., 2003; Moyne et al., 2001). The antimicrobial activity exhibited by Bacillus sp. is dependent on the culture medium composition, and different nitro- gen sources can result in the production of different antibiotics (Besson et al., 1987; Chevanet et al., 1986; Davis et al., 1999; Volpon et al., 2000). Aspartic acid is the preferred nitrogen source for the production of iturinic compounds by Bacillus subtilis (Besson et al., 1987) and Bacillus amyloliquefaciens (Caldeira et al., 2006, 2007, 2008). With increasing culture time, the nutrient content changes and adverse environmental conditions appear. Thus, incubation time is another factor influencing antibiotic production (Caldeira et al., 2008; Feio et al., 2004; Moyne et al., 2001), as the response to adverse environmental conditions can lead to activation of differ- ent mechanisms for the production of antibiotics giving a compet- itive advantage to the producer microorganism (Dieckmann et al., 2001). The link between antibiotic production and Bacillus sporulation is not fully understood. During production of lipopeptides in sub- ⇑ Corresponding author. Tel.: +351 266 745 315; fax: +351 266 745 303. E-mail address: hvicente@uevora.pt (H. Vicente). 0960-8524/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.biortech.2010.07.080 abstract The combined effect of incubation time (IT) and aspartic acid concentration (AA) on the predicted bio- mass concentration (BC), Bacillus sporulation (BS) and anti-fungal activity of compounds (AFA) produced by Bacillus amyloliquefaciens CCMI 1051, was studied using Artificial Neural Networks (ANNs). The values predicted by ANN were in good agreement with experimental results, and were better than those obtained when using Response Surface Methodology. The database used to train and validate ANNs con- tains experimental data of B. amyloliquefaciens cultures (AFA, BS and BC) with different incubation times (1–9 days) using aspartic acid (3–42 mM) as nitrogen source. After the training and validation stages, the 2–7-6–3 neural network results showed that maximum AFA can be achieved with 19.5 mM AA on day 9; however, maximum AFA can also be obtained with an incubation time as short as 6 days with 36.6 mM AA. Furthermore, the model results showed two distinct behaviors for AFA, depending on IT.