Degradation of Poly(Butylene Succinate) and Poly(Butylene Succinate)/Poly(Lactide) Blends using Serine Protease Produced from Laceyella sacchari LP175

The thermophilic filamentous bacterium Laceyella sacchari LP175 was cultivated in a 10.0 L airlift fermenter to produce serine protease at 50 °C. Maximal serine protease activity at 1,123.32 ± 15.8 U/mL was obtained for cultivation at 0.6 vvm aeration rate for 36 h. The crude enzyme was applied for degradation of poly (butylene succinate) (PBS), and poly (butylene succinate)/poly(lactide) blend (PBS/PLA) powders at 50 °C for 48 h with different substrates and enzyme concentrations. Results showed that serine protease produced from L. sacchari LP175 degraded PBS and PBS/PLA at 46.5 ± 2.05 and 49.8 ± 1.45 %, respectively, at an initial substrate concentration of 100 g/L with 1,200 U/mL of serine protease activity. Percentage degradation of PBS and PBS/PLA was improved to 51.4 ± 1.06 and 56.9 ± 1.42 %, respectively, when upscaled in a 2.0 L stirrer fermenter with 200 rpm agitation rate. Degradation products evaluated by a scanning electron microscope (SEM) and Fourier transform infrared spectroscopy (FTIR) confirmed that serine protease produced from L. sacchari LP175 degraded both PBS and PBS/PLA polymers. Results showed that microbial enzyme technology could be used to degrade PBS and PBS/PLA blend polymers and reduce the accumulation of waste. HIGHLIGHTS Upscaled serine protease production was achieved in a 10 L airlift fermenter by sacchari LP175 using low-cost agricultural products as substrate The crude enzyme degraded PBS and PBS/PLA powders (100 g/L) at up to 51.4 and 56.9 %, respectively in a 2.0 L stirrer fermenter under optimal conditions Degradation products of PBS and PBS/PLA by crude enzyme produced from sacchari LP175 were characterized GRAPHICAL ABSTRACT

Kinetics of Crude Peroxidase from the Rind of Watermelon Fruit

Aims: To study the kinetics of crude peroxidase from the rind of watermelon fruit in various assay conditions. Study Design: In vitro enzyme assay. Place and Duration of Study: Department of Biochemistry, Faculty of Life Sciences, Ambrose Alli University, Ekpoma, Edo State, Nigeria between October 2015 and January 2016. Methodology: The activity of the crude peroxidase extracted from the rind of watermelon was determined by measuring the rate of oxidation of KI at 25oC in a 3.0 ml reaction mixture which contained 2.3 ml of 25 mM - 400 mM sodium acetate buffer (pH 3.5-6.0), 0.2 ml of 2 mM KI, 0.1ml of the crude peroxidase, and 0.2 ml of varying concentrations of chlorpromazine (0.01 mM - 0.1 mM). In all cases, 0.2 ml of 0.01 mM – 1 mM H2O2 was added last to initiate the reaction. Only one parameter was varied per assay. Assays were done in five replicates. The initial velocity of the crude peroxidase for KI oxidation was determined using the absorbance at 353 nm. Results: The concentration of H2O2 that generated an optimal activity for the crude peroxidase extracted was 0.2 mM, while a pH of 5.5 was optimal for the crude enzyme. The activity of the crude enzyme increased proportionately within a buffer concentration range of 25 mM and 400 mM. Chlorpromazine (0.01 mM - 0.1 mM) proportionately increased the enzyme activity, while promethazine within a range of 0.01 mM and 0.06 mM proportionally increased the enzyme activity. Further increase in promethazine concentration beyond 0.6 mM resulted in a decreased activity of the enzyme. Conclusion: This study suggests that the Rind of watermelon is an alternative source of peroxidase. The activity of this peroxidase can be enhanced by high buffer concentrations in the presence of some redox mediators like promethazine and chlorpromazine at a pH of 5.5.

Kinetic Analysis of Crude Enzyme Extract Produced Via Solid State Fermentation of Banana Pseudo Stem Waste

Banana Pseudo stem waste after each harvest contributes about 70–80 Milli Tons Per hector. The banana pseudo stem will be thrown as waste biomass after each harvest as it is unstable for the upcoming harvest. The biggest challenge in banana cultivation is the utilization of biomass of banana pseusostem waste into valuable products. In this study, Xylano-pectinase enzyme extract was produced from banana pseudo stem waste under solid-state fermentation by Enterobacter cloacae PMC04. The highest pectinase and xylanase activities obtained using banana pseudo stem as carbon source were 124.62 U/ml and 173.81 U/ml respectively. Thermodynamics stated that range 40-50oC were considered to be the optimal temperature for xylano-pectinase enzyme production and subsequent degumming of banana fibers. The crude enzyme extract were then used in the degumming of banana fibers for textile application. Textile processing of banana fiber necessitates the removal of hemicellulose substance which can be achieved by crude xylano-pectinase enzyme. It was found that crude xylano-pectinase was efficient in the removal of hemicellulose substance from the fibers. Results obtained from this study demonstrate that the proposed bioprocess could be successfully applied for the degumming of banana fibers sustainably.

Proteases Production and Chitin Preparation from the Liquid Fermentation of Chitinous Fishery By-Products by Paenibacillus elgii

Chitinous fishery by-products have great application in the production of various bioactive compounds. In this study, Paenibacillus elgii TKU051, a protease-producing bacterial strain, was isolated using a medium containing 1% squid pens powder (SPP) as the sole carbon/nitrogen (C/N) source. P. elgii TKU051 was found to produce at least four proteases with molecular weights of 100 kDa, 57 kDa, 43 kDa, and 34 kDa (determined by the gelatin zymography method). A P. elgii TkU051 crude enzyme cocktail was optimally active at pH 6–7 and 60 °C. The 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity and α-glucosidase inhibitory activity of the hydrolysates obtained from the hydrolysis of shrimp shell powder, shrimp head powder, shrimp meat powder, fish head powder and soya bean powder catalyzed by the P. elgii TkU051 crude enzyme cocktail were also evaluated. P. elgii TKU051 exhibited a high deproteinization capacity (over 94%) on different kinds of shrimp waste (shrimp heads and shells; fresh and cooked shrimp waste; shrimp waste dried by oven and lyophilizer), and the Fourier-transform infrared spectroscopy profile of the chitin obtained from the deproteinization process displayed the characteristic of chitin. Finally, the obtained chitin exhibited an effect comparable to commercial chitin in terms of adsorption against Congo Red (90.48% and 90.91%, respectively). Thus, P. elgii TKU051 showed potential in the reclamation of chitinous fishery by-products for proteases production and chitin extraction.

Safety Evaluation and Whole Genome Sequencing of Aspergillus japonicas PJ01 Reveal Its Potential to Degrade Citrus Segments in Juice Processing

Aspergillus japonicas PJ01 (A. japonicas PJ01) is a strain isolated from the rotten branches. In previ-ous studies, it was shown that it can produce complex enzymes to degrade polysaccharide com-ponents. In this study, we evaluated the safety of its crude enzyme solution. Acute oral toxicity, subchronic toxicity, micronucleus and sperm malformation tests all validated the high biologi-cal safety for the crude enzymes. Secondly, we carried out the citrus segment degradation ex-periment of crude enzyme solution. Compared with the control group, the crude enzyme solu-tion of A. japonicas PJ01 can completely degrade the segments in 50 min, which provides the basis for enzymatic peeling during juice processing. The whole genome sequencing showed that the genome of A. japonicus PJ01 has a GC content of 51.37% with a size of 36204647 bp, and encoded 10070 genes. GO, COG, KEGG and CAZy databases were used in gene annotation analyses. Pathway enrichment showed many genes related to carbohydrate metabolism, rich in genes re-lated to pectinase, xylanase and carboxylcellulase. Therefore, the complex enzyme produced by A. japonicus PJ01 can be used in gizzard juice processing to achieve efficient enzymatic decapsu-lation.

IN VITRO STUDY ON PROTEASE AND THROMBOLYTIC ACTIVITY OF AQUEOUS EXTRACT FROM LEUCAS ASPERA (L.) LEAVES

Objective: The current study is an attempt to screen for the in vitro clot lysis and proteolytic activity of aqueous extract of Leucas aspera leaves. Methods: Thrombolytic activity and protease activity of the crude enzyme obtained by ammonium sulfate precipitation and dialysis were assayed using blood clot and casein as substrates respectively. Native PAGE and gel documentation studies were performed to calculate the molecular weight of the enzyme. Results: In the study, 40% salt fractioned crude enzyme sample exhibited significant thrombolytic and caseinolytic activity. Further dose-dependent increased activity was observed with the maximum lytic activity of 52.11±1.04 % at 1 mg/ml of the sample when compared to the reference drug streptokinase (71.39±0.32%). Also, 68.72±0.62 U/hr of caseinolytic activity was observed for 1 mg/ml of the sample fraction. Conclusion: The study highlights and validates the efficacy of Leucas aspera leaves extract for thrombolytic and proteolytic actions. Enzyme with an approximate molecular weight, 19.89 KDa could be responsible for the significant lytic activity.