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An aqueous protocol was developed for nattokinase purification under scalable conditions using a combination of salting out, anion exchange chromatography and ultra-filtration. The enzyme was firstly washed off from the fermented soybeans. The whole proteins were precipitated by ammonium sulfate and purified by anion exchange chromatography. The purified enzyme was finally refined by size exclusion chromatography and ultra-filtration at laboratory and industry-compatible level, respectively. There was a 476.18-fold increase in enzymatic activity with a 48.3% yield at the laboratory level and 429.75-fold increase in enzymatic activity with a 42.7% yield at the industrial-compatible scale. The protocol was universally adaptive for purifying the fibrinolytic enzymes produced by bacillus tequilensis, bacillus amyloliquefaciens, and bacillus cereus, showing a purification efficiency of 329.76-fold with 42.7% yield, 221.78-fold with 32.5% yield, and 288.56-fold with 38.7% yield, respectively. All procedures are scalable, aqueous, simple, cost-effective, and suitable for fibrinolytic enzymes industrial production.

Aims: Nonalcoholic fatty liver disease (NAFLD) is a chronic liver disease that affects adipose function. This study aimed to explore the function of adipocytes-derived exosomal (ADEs) miR-122 in NAFLD., Methods: A high-fat and high-fructose diet-induced rat model and a palmitic acid (PA)-induced in vitro model were established. The RNA level of miR-122 and Sirt1 was measured using qRT-PCR. The protein levels of exosome biomarkers, and lipogenesis, inflammation and fibrosis biomarkers were determined by western blotting. Cell viability and apoptosis were assessed using cell counting kit-8 and flow cytometry, respectively. Serum alanine aminotransferase, aspartate aminotransferase, total cholesterol, triglyceride levels were measured. Liver tissue damage was assessed using haematoxylin and eosin staining. The interaction between miR-122 and Sirt1 3'UTR was assessed using a luciferase reporter gene assay., Results: ADEs exhibited abundant level of miR-122 and promoted lipogenesis, impaired hepatocyte survival, enhanced liver damage and increased serum lipid levels in vivo and in vitro. Inhibition of miR-122 in ADEs alleviated NAFLD progression, lipid and glucose metabolism, liver inflammation and fibrosis both in vivo and in vitro. miR-122 binds directly to the 3'UTR of Sirt1 to suppress its expression. Moreover, Sirt1 overexpression reversed the increase in cell apoptosis, glucose and lipid metabolism, liver inflammation and fibrosis induced by ADEs in vivo and in vitro., Conclusions: The ADEs miR-122 promotes the progression of NAFLD via modulating Sirt1 signalling in vivo and in vitro. The ADEs miR-122 may be a promising diagnostic biomarker and therapeutic target for NAFLD., (Copyright © 2022 Elsevier España, S.L.U. All rights reserved.)