An extracellular lipase from Aureobasidium pullulans was obtained and purified with a specific activity of 17.7 U/mg of protein using ultrafiltration and a DEAE-Sepharose Fast Flow column. Characterization of the lipase indicated that it is a novel finding from the species A. pullulans. The molecular weight of the lipase was 39.5 kDa, determined by sodium dodecyl sulfonate-polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme exhibited its optimum activity at 40 °C and pH of 7. It also showed a remarkable stability in some organic solutions (30%, v/v) including n-propanol, isopropanol, dimethyl sulfoxide (DMSO), and hexane. The catalytic activity of the lipase was enhanced by Ca²⁺ and was slightly inhibited by Mn²⁺ and Zn²⁺ at a concentration of 10 mmol/L. The lipase was activated by the anionic surfactant SDS and the non-ionic surfactants Tween 20, Tween 80, and Triton X-100, but it was drastically inhibited by the cationic surfactant cetyl trimethyl ammonium bromide (CTAB). Furthermore, the lipase was able to hydrolyze a wide variety of edible oils, such as peanut oil, corn oil, sunflower seed oil, sesame oil, and olive oil. Our study indicated that the lipase we obtained is a potential biocatalyst for industrial use.
Ascomycota/enzymology* ; Calcium ; Catalysis ; Corn Oil/metabolism* ; Detergents/chemistry* ; Enzyme Stability ; Fungal Proteins/chemistry* ; Glucans/chemistry* ; Hexanes/chemistry* ; Hydrogen-Ion Concentration ; Hydrolysis ; Industrial Microbiology ; Lipase/chemistry* ; Manganese/chemistry* ; Olive Oil/metabolism* ; Peanut Oil/metabolism* ; Sesame Oil/metabolism* ; Substrate Specificity ; Sunflower Oil/metabolism* ; Surface-Active Agents ; Temperature ; Zinc/chemistry*

OBJECTIVE: To investigate the effect of curcumol on NOD-like receptor thermoprotein domain 3 (NLRP3) inflammasomes, and analyze the mechanism underlying curcumol against liver fibrosis. METHODS: Thirty Kunming mice were divided into a control group, a model group and a curcumol group according to a random number table, 10 mice in each group. Mice were intraperitoneally injected with 40% carbon tetrachloride (CCl₄:peanut oil, 2:3 preparation) at 5 mL/kg for 6 weeks, twice a week, for developing a liver fibrosis model. The mice in the control group were given the same amount of peanut oil twice a week for 6 weeks. The mice in the curcumol group were given curcumol (30 mL/kg) intragastrically, and the mice in the model and control groups were given the same amount of normal saline once a day for 6 weeks. Changes in liver structure were observed by hematoxylin and eosin (HE) and Masson staining. Liver function, liver fiber indices, and the expression of interleukin (IL)-10 and tumor necrosis factor-α (TNF-α) levels were determined by automatic biochemical analyzer and enzyme linked immunosorbent assay kit. Immunoblotting and reverse transcription-quantitative PCR (RT-qPCR) were performed to detect the expression of NLRP3 inflammasome-related molecules, TGF-β and collagen. RESULTS: HE and Masson staining results showed that the hepatocytes of the model group were arranged irregularly with pseudo-lobular structure and a large amount of collagen deposition. The mice in the curcumol group had a significant decrease in liver function and liver fibers indices compared with the model group (P<0.05); RT-qPCR and Western blotting results reveal that, in the curcumol group, the mRNA and protein expression levels of NLRP3, IL-1 β, Caspase 1 and gasdermin D decreased significantly compared with the model group (P<0.05); immunohistochemical results showed that in the curcumol group, the protein expression levels of NLRP3 and IL-1 β decreased significantly compared with the model group (P<0.05). CONCLUSION A potential anti-liver fibrosis mechanism of curcumol may be associated with the inhibition of NLRP3 inflammasomes and decreasing the downstream inflammatory response.
Animals ; Mice ; Inflammasomes/metabolism* ; Interleukin-1beta/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; NLR Proteins ; Caspase 1 ; Tumor Necrosis Factor-alpha ; Carbon Tetrachloride ; Hematoxylin ; Saline Solution ; Eosine Yellowish-(YS) ; Peanut Oil ; Liver Cirrhosis/drug therapy* ; RNA, Messenger/genetics* ; Collagen ; Transforming Growth Factor beta