Pilot-scale production of Bacillus subtilis MSCL 897 spore biomass and antifungal secondary metabolites in a low-cost medium.

OBJECTIVES: Bacillus subtilis is a plant growth promoting bacterium (PGPB) that acts as a microbial fertilizer and biocontrol agent, providing benefits such as boosting crop productivity and improving nutrient content. It is able to produce secondary metabolites and endospores simultaneously, enhancing its ability to survive in unfavorable conditions and eliminate competing microorganisms. Optimizing cultivation methods to produce B. subtilis MSCL 897 spores on an industrial scale, requires a suitable medium, typically made from food industry by-products, and optimal temperature and pH levels to achieve high vegetative cell and spore densities with maximum productivity. RESULTS: This research demonstrates successful pilot-scale (100 L bioreactor) production of a biocontrol agent B. subtilis with good spore yields (1.5 × 109 spores mL-1) and a high degree of sporulation (>80%) using a low-cost cultivation medium. Culture samples showed excellent antifungal activity (1.6-2.3 cm) against several phytopathogenic fungi. An improved methodology for inoculum preparation was investigated to ensure an optimal seed culture state prior to inoculation, promoting process batch-to-batch repeatability. Increasing the molasses concentration in the medium and operating the process in fed-batch mode with additional molasses feed, did not improve the overall spore yield, hence, process operation in batch mode with 10 g molasses L-1 is preferred. Results also showed that the product quality was not significantly impacted for up to 12 months of storage at room temperature. CONCLUSION: An economically-feasible process for B. subtilis-based biocontrol agent production was successfully developed at the pilot scale.

Butter whey and corn steep liquor as sole raw materials to obtain a bioemulsifier from Yarrowia lipolytica for food oil-in-water emulsions / Soro de manteiga e milhocina como únicas matérias primas para obter bioemulsificante de Yarrowia lipolytica para emulsões óleo-em-água alimentícias

ABSTRACT: A synthetic medium containing glucose, glycerol, yeast extract (YE), and ammonium sulfate (AS) was compared to several low-cost media in their ability to produce high emulsification index (EI). The goal was to reduce the production costs of an emulsifier with application in food oil-in-water emulsions. To this end, agro-industrial by-products were screened for bioemulsifier production from Yarrowia lipolytica. The statistical analysis showed that the EIs of media containing residual frying oil from palm oil (RFO_palm) or soybean oil (RFO_soy), residual liquid from butter production (butter whey, BWhey) or cheese production (cheese whey, CWhey), supplemented with YE and AS were similar to the EI of the synthetic medium. The replacement of YE by corn steep liquor (CSL) also resulted in similar EI, except for RFO_soy. BWhey was tested with CSL without AS and similar EI (66.8%) was detected in comparison to that of the same medium with AS (66.3%). The cell-free broth obtained after Y. lipolytica growth in BWhey+CSL was successfully used to obtain vegetable oil-in-water emulsions indicating its potential application in food products.

RESUMO: O meio sintético contendo glicose, glicerol, extrato de levedura (YE) e sulfato de amônio (AS) foi comparado a meios de baixo custo para produzir alto índice de emulsificação (EI). Para este fim, os subprodutos agroindustriais foram rastreados quanto à produção de bioemulsificante por Yarrowia lipolytica. A análise estatística mostrou que EI de meios contendo óleo de fritura residual de óleo de palma (RFO_palm) ou óleo de soja (RFO_soy), líquido residual da produção de manteiga (soro de manteiga, BWhey) ou da produção de queijo (soro de queijo, CWhey), suplementado com YE e AS foram semelhantes ao EI do meio sintético. A substituição do YE por milhocina (CSL) também resultou em EI semelhante, exceto no RFO_soy. O BWhey foi testado com CSL sem AS e EI semelhante (66,8%) foi detectado em comparação com o mesmo meio com AS (66,3%). O meio isento de células obtido após o crescimento de Y. lipolytica no meio BWhey + CSL foi utilizado com sucesso para obter emulsões de óleo vegetal em água, indicando sua potencial aplicação em produtos alimentícios.

Utilizing Gelatinized Starchy Waste from Rice Noodle Factory as Substrate for L(+)-Lactic Acid Production by Amylolytic Lactic Acid Bacterium Enterococcus faecium K-1.

To valorize starchy waste from rice noodle factory, bioconversion of gelatinized starchy waste (GSW) to value-added product as L(+)-lactic acid, the monomer for polylactate synthesis, was investigated using amylolytic lactic acid bacterium, Enterococcus faecium K-1. Screening for appropriate nitrogen source to replace expensive organic nitrogen sources revealed that corn steep liquor (CSL) was the most suitable regarding high efficacy for L(+)-LA achievement and low-cost property. The successful applying statistic experimental design, Plackett-Burman design incorporated with central composite design (CCD), predicted the maximum L(+)-LA of 93.07 g/L from the optimized medium (OM) containing 125.7 g/L GSW and 207.3 g/L CSL supplemented with CH3COONa, MgSO4, MnSO4, K2HPO4, CaCl2, (NH4)2HC6H5O7, and Tween80. Minimizing the medium cost by removal of all inorganic salts and Tween80 from OM was not an effect on L(+)-LA yield. Fermentation using the optimized medium without minerals (OM-Mi) containing only GSW (125.7 g/L) and CSL (207.3 g/L) in a 10-L fermenter was also successful. Thinning GSW with α-amylase from Lactobacillus plantarum S21 increased L(+)-LA productivity in the early stage of 24-h fermentation. Not only showing the feasible bioconversion process for GSW utilizing as a substrate for L(+)-LA production, this research also demonstrated the efficient model for industrial starchy waste valorization.

Screening and optimization of low-cost medium for Pseudomonas putida Rs-198 culture using RSM

The plant growth-promoting rhizobacterial strain Pseudomonas putida Rs-198 was isolated from salinized soils from Xinjiang Province. We optimized the composition of the low-cost medium of P. putida Rs-198 based on its bacterial concentration, as well as its phosphate-dissolving and indole acetic acid (IAA)-producing capabilities using the response surface methodology (RSM), and a mathematical model was developed to show the effect of each medium component and its interactions on phosphate dissolution and IAA production. The model predicted a maximum phosphate concentration in medium containing 63.23 mg/L inorganic phosphate with 49.22 g/L corn flour, 14.63 g/L soybean meal, 2.03 g/L K2HPO4, 0.19 g/L MnSO4 and 5.00 g/L NaCl. The maximum IAA concentration (18.73 mg/L) was predicted in medium containing 52.41 g/L corn flour, 15.82 g/L soybean meal, 2.40 g/L K2HPO4, 0.17 g/L MnSO4 and 5.00 g/L NaCl. These predicted values were also verified through experiments, with a cell density of 10(13) cfu/mL, phosphate dissolution of 64.33 mg/L, and IAA concentration of 18.08 mg/L. The excellent correlation between predicted and measured values of each model justifies the validity of both the response models. The study aims to provide a basis for industrialized fermentation using P. putida Rs-198.

Screening and optimization of low-cost medium for Pseudomonas putida Rs-198 culture using RSM.

The plant growth-promoting rhizobacterial strain Pseudomonas putida Rs-198 was isolated from salinized soils from Xinjiang Province. We optimized the composition of the low-cost medium of P. putida Rs-198 based on its bacterial concentration, as well as its phosphate-dissolving and indole acetic acid (IAA)-producing capabilities using the response surface methodology (RSM), and a mathematical model was developed to show the effect of each medium component and its interactions on phosphate dissolution and IAA production. The model predicted a maximum phosphate concentration in medium containing 63.23 mg/L inorganic phosphate with 49.22 g/L corn flour, 14.63 g/L soybean meal, 2.03 g/L K2HPO4, 0.19 g/L MnSO4 and 5.00 g/L NaCl. The maximum IAA concentration (18.73 mg/L) was predicted in medium containing 52.41 g/L corn flour, 15.82 g/L soybean meal, 2.40 g/L K2HPO4, 0.17 g/L MnSO4 and 5.00 g/L NaCl. These predicted values were also verified through experiments, with a cell density of 10(13) cfu/mL, phosphate dissolution of 64.33 mg/L, and IAA concentration of 18.08 mg/L. The excellent correlation between predicted and measured values of each model justifies the validity of both the response models. The study aims to provide a basis for industrialized fermentation using P. putida Rs-198.