Improved production of D-pantothenic acid in Escherichia coli by integrated strain engineering and fermentation strategies.

D-pantothenic acid (D-PA) is an essential vitamin that has been widely used in medicine, food, and animal feed. Microbial production of D-PA from natural renewable resources is attractive and challenging. In this study, both strain improvements and fermentation process strategies were applied to achieve high-level D-PA production in Escherichia coli. First, a D-PA-producing strain was developed through deletion of the aceF and mdh genes combined with the overexpression of the gene ppnk. The obtained engineered E. coli DPA02/pT-ppnk accumulated 6.89 ± 0.11 g/L of D-PA in shake flask fermentation, which was 79.9 % higher than the control strain. Moreover, the cultivation process contributed greatly to D-PA production with respect to titer and productivity by betaine supplementation and dissolved oxygen (DO)-feedback feeding framework. Under optimal conditions, 68.3 g/L of D-PA, the specific productivity of 0.794 g/L h and the yield of 0.36 g/g glucose in 5 L fermenter were achieved. Overall, this research successfully exploited advanced strategies to lay the foundation for bio-based D-PA production in industrial applications.

Overproduction of D-pantothenic acid via fermentation conditions optimization and isoleucine feeding from recombinant Escherichia coli W3110.

D-pantothenic acid (D-PA), as a crucial vitamin, is widely used in food, animal feed, cosmetics, and pharmaceutical industries. In our previous work, recombinant Escherichia coli W3110 for production of D-PA was constructed through metabolic pathway modification. In this study, to enhance D-PA production, statistical optimization techniques including Plackett-Burman (PB) design and Box-Behnken design (BBD) first were adopted to optimize the culture condition. The results showed that the glucose, ß-alanine and (NH4)2SO4 have the most significant effects on D-PA biosynthesis. The response surface model based on BBD predicted that the optimal concentration is glucose 56.0 g/L, ß-alanine 2.25 g/L and (NH4)2SO4 11.8 g/L, the D-PA titer increases from 3.2 g/L to 6.73 g/L shake flask fermentation. For the fed-batch fermentation in 5 L fermenter, the isoleucine feeding strategy greatly increased the titer and productivity of D-PA. As a result, titer (31.6 g/L) and productivity (13.2 g/L·d) of D-PA were achieved, they increased by 4.66 times and 2.65 times, respectively, compared with batch culture. At the same time, the accumulation of acetate reduced from 29.79 g/L to 8.55 g/L in the fed-batch fermentation. These results demonstrated that the optimization of medium composition and the cell growth rate are important to increase the concentration of D-PA for microbial fermentation. This work laid the foundation for further research on the application of D-PA microbial synthesis. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02773-0.

Insecticidal activities of constituents of Litsea cubeba fruit extracts effective against the maize weevil (Coleoptera: Curculionidae).

In this study, we investigated the insecticidal activities, including contact toxicity, fumigant toxicity, and repellent activity, of Litsea cubeba fruit extracts against Sitophilus zeamais Motschulsky (Coleoptera: Curculionidae). The extracts, obtained by liquid-liquid extraction in n-hexane, ethyl acetate, chloroform, and water were analyzed by gas chromatography-mass spectrometry. Among the different extract types, chloroform extracts exhibited the strongest repellent, contact, and fumigant activities against S. zeamais. The main components of the chloroform extracts were identified as laurine (21.15%) and 2,6-diisopropyl aniline (16.14%), followed by chlorobutanol (10.54%), 3-O-methyl-N-acetyl-d-glucosamine (10.03%), and 6-methyl-5-hepten-2-one (8.33%). Among the identified components of the chloroform extracts, chlorobutanol showed the strongest fumigant toxicity (LD50 = 21.91 mg/liter), contact toxicity (LD50 = 54.25 µg/adult), and repellent activity against S. zeamais. These results indicate that L. cubeba fruit extracts possess natural insecticide-like activities against S. zeamais.