Mechanism and evolutionary analysis of Yarrowia lipolytica CA20 capable of producing erythritol with a high yield based on comparative genomics.

Combined mutagenesis is widely applied for the breeding of robust Yarrowia lipolytica used in the production of erythritol. However, the changes of genome after mutagenesis remains unclear. This study aimed to unravel the mechanism involved in the improved erythritol synthesis of CA20 and the evolutionary relationship between different Y. lipolytica by comparative genomics analysis. The results showed that the genome size of Y. lipolytica CA20 was 20,420,510 bp, with a GC content of 48.97%. There were 6330 CDS and 649 ncRNA (non-coding RNA) in CA20 genome. Average nucleotide identity (ANI) analysis showed that CA20 genome possessed high similarity (ANI > 99.50%) with other Y. lipolytica strains, while phylogenetic analysis displayed that CA20 was classified together with Y. lipolytica IBT 446 and Y. lipolytica H222. CA20 shared 5342 core orthologous genes with the 8 strains while harbored 65 specific genes that mainly participated in the substrate and protein transport processes. CA20 contained 166 genes coding for carbohydrate-active enzymes (CAZymes), which was more than that found in other strains (108－137). Notably, 4, 2, and 13 different enzymes belonging to glycoside hydrolases (GHs), glycosyltransferases (GTs), and carbohydrate esterases (CEs), respectively, were only found in CA20. The enzymes involved in the metabolic pathway of erythritol were highly conserved in Y. lipolytica, except for transaldolase (TAL1). In addition, the titer and productivity of erythritol by CA20 were 190.97 g/L and 1.33 g/L/h, respectively, which were significantly higher than that of WT5 wherein 128.61 g/L and 0.92 g/L/h were obtained (P< 0.001). Five frameshift mutation genes and 15 genes harboring nonsynonymous mutation were found in CA20 compared with that of WT5. Most of these genes were involved in the cell division, cell wall synthesis, protein synthesis, and protein homeostasis maintenance. These findings suggested that the genome of Y. lipolytica is conserved during evolution, and the variance of living environment is one important factor leading to genome divergence. The varied number of CAZymes existed in Y. lipolytica is one factor that contributes to the performance difference. The increased synthesis of erythritol by Y. lipolytica CA20 is correlated with the improvement of the stability of cell structure and internal environment. The results of this study provide a basis for the directional breeding of robust strains used in erythritol production.

Enrichment and separation of steroidal saponins from the fibrous roots of Ophiopogon japonicus using macroporous adsorption resins.

In this study, a simple and effective strategy for the enrichment of total steroidal saponins (TSS) from the fibrous roots of Ophiopogon japonicus (L. f.) Ker-Gawl. (FROJ) using macroporous adsorption resin was systematically developed. XAD-7HP resin was selected from six macroporous resins for further study because of the highest static adsorption and desorption capacities. The static adsorption of TSS on XAD-7HP resin fitted well to the Langmuir isotherm model and pseudo second-order kinetic model; the thermodynamics test showed that the adsorption process was spontaneous and exothermic. The dynamic tests on XAD-7HP resin columns demonstrated that the breakthrough volume was 16 bed volume (BV), and 6 BV of 80% ethanol was suitable for dynamic desorption. In a lab scale-up separation under optimal dynamic conditions, the content of TSS in the resin-enrichment fraction increased from 1.83% in the crude extracts to 13.86% by 7.59-fold with a recovery yield of 82.68%. Three steroidal saponins were obtained from the resin-enrichment fraction, and showed protective effects against oxidized low-density lipoprotein (ox-LDL) induced human umbilical vein endothelial cell (HUVEC) injury. Overall, these results suggested that XAD-7HP resin chromatography was an effective strategy for the large scale enrichment of TSS from FROJ, which showed the potential for functional food and pharmaceutical application.

Biotreatment of oily wastewater by rhamnolipids in aerated active sludge system.

Oily wastewater generated by various industries creates a major ecological problem throughout the world. The traditional methods for the oily wastewater treatment are inefficient and costly. Surfactants can promote the biodegradation of petroleum hydrocarbons by dispersing oil into aqueous environment. In the present study, we applied rhamnolipid-containing cell-free culture broth to enhance the biodegradation of crude oil and lubricating oil in a conventional aerobically-activated sludge system. At 20 degrees C, rhamnolipids (11.2 mg/L) increased the removal efficiency of crude oil from 17.7% (in the absence of rhamnolipids) to 63%. At 25 degrees C, the removal efficiency of crude oil was over 80% with the presence of rhamnolipids compared with 22.3% in the absence of rhamnolipids. Similarly, rhamnolipid treatment (22.5 mg/L) for 24 h at 20 degrees C significantly increased the removal rate of lubricating oil to 92% compared with 24% in the absence of rhamnolipids. The enhanced removal of hydrocarbons was mainly attributed to the improved solubility and the reduced interfacial tension by rhamnolipids. We conclude that a direct application of the crude rhamnolipid solution from cell culture is effective and economic in removing oily contaminants from wastewater.