Valorization of whey-based side streams for microbial biomass, molecular hydrogen, and hydrogenase production.

Side streams of the dairy industry are a suitable nutrient source for cultivating microorganisms, producing enzymes, and high-value chemical compounds. The heterotrophic Escherichia coli and chemolithoautotroph Ralstonia eutropha are of major biotechnological interest. R. eutropha is a model organism for producing O2-tolerant [NiFe]-hydrogenases (Hyds) (biocatalysts), and E. coli has found widespread use as an expression platform for producing recombinant proteins, molecular hydrogen (H2), and other valuable products. Aiming at developing suitable cultivation media from side streams of the dairy industry, the pre-treatment (filtration, dilution, and pH adjustment) of cheese (sweet) whey (SW) and curd (acid) whey (AW), with and without the use of ß-glucosidase, has been performed. Growth parameters (oxidation-reduction potential (ORP), pH changes, specific growth rate, biomass formation) of E. coli BW25113 and R. eutropha H16 type strains were monitored during cultivation on filtered and non-filtered SW and AW at 37 °C, pH 7.5 and 30 °C, pH 7.0, respectively. Along with microbial growth, measurements of pH and ORP indicated good fermentative growth. Compared to growth on fructose-nitrogen minimal salt medium (control), a maximum cell yield (OD600 4.0) and H2-oxidizing Hyd activity were achieved in the stationary growth phase for R. eutropha. Hyd-3-dependent H2 production by E. coli utilizing whey as a growth substrate was demonstrated. Moreover, good biomass production and prolonged H2 yields of ~ 5 mmol/L and cumulative H2 ~ 94 mL g/L dry whey (DW) (ß-glucosidase-treated) were observed during the cultivation of the engineered E. coli strain. These results open new avenues for effective whey treatment using thermostable ß-glucosidase and confirm whey as an economically viable commodity for biomass and biocatalyst production. KEY POINTS: • Archaeal thermostable ß-glucosidase isolated from the metagenome of a hydrothermal spring was used for lactose hydrolysis in whey. • Hydrogenase enzyme activity was induced during the growth of Ralstonia eutropha H16 on whey. • Enhanced biomass and H2 production was shown in a genetically modified strain of Escherichia coli.

Synthesis, Characterization, and Study of Catalytic Activity of Chiral Cu(II) and Ni(II) Salen Complexes in the α-Amino Acid C-α Alkylation Reaction.

A new family of Cu(II) and Ni(II) salen complexes was synthesized and fully characterized through various physicochemical methods. Their catalytic activity was evaluated in the phase transfer Cα-alkylation reaction of the Schiff bases of D,L-alanine ester and benzaldehyde derivatives. It was found that the introduction of a chlorine atom into the ortho- and para-positions of the phenyl ring of the substrate resulted in an increase in both the chemical yield and the asymmetric induction (ee 66-98%). The highest enantiomeric excess was achieved in the case of a Cu(II) salen complex based on (S,S)-cyclohexanediamine and salicylaldehyde at -20 °C. The occurrence of a bulky substituent in the ligand present in the complexes led to a drastic decrease in ee and chemical yield. For instance, the introduction of bulky substituents at positions 3 and 5 of the phenyl ring of the catalyst resulted in a complete loss of the stereoselectivity control in the alkylation reaction.