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Silkworm pupae are a major by-product of the silkworm mulberry industry, and the high content of tri-alpha-linolenin (glyceryl tri-alpha-linolenate, LnLnLn) in silkworm pupae oil makes it an excellent essential fatty acid supplement. However, the complex triglyceride composition of silkworm pupae oil limits its absorption and utilization by the human body. In this study, a sequential simulated moving bed (SSMB) approach for continuous separation was presented to recover the high-value tri-alpha-linolenin monomer from silkworm pupae oil. C18 particles were used as the stationary phase. Acetonitrile and isopropanol were used as the elution mobile phases. Under optimal SSMB conditions, the purity and yield of the extracted products were 95.15±2.06% and 2.14±0.06 mg/mL, respectively. In comparison to the conventional extraction approach, the yield of tri-alpha-linolenin separated by SSMB was 26.7 times greater, the dose of eluent was lowered by 2 times, and the eluent could be recycled. To validate the experimental parameters, a mass transfer diffusion model was used. The molecular diffusion coefficient of tri-alpha-linolenin was 2.61×10−6 cm2/s, the axial diffusion coefficient was 2.61×10−6+7.56×10−3μ, and the mass transfer diffusion coefficient was 0.87 s−1. The correlation coefficient between the model and the actual measured values was 0.9688. Consequently, the separation of tri-alpha-linolenin monomer in silkworm pupae oil by sequential simulated moving bed chromatography column can be reasonably described and predicted by this model. In conclusion, a novel approach to the commercialization of insect oils was developed by the extraction and purification of tri-alpha-linolenin from silkworm pupae oil using the SSMB approach. [ABSTRACT FROM AUTHOR]

The present study aimed at developing chitosan/collagen protease-based composite membranes loaded with enzyme nanoparticles and accessing its efficient biodegradation using a recombinantly produced aspartic protease from Aspergillus fumigatus (AfAP) with functional biochemical properties. By using Fourier transform infrared spectroscopy (FT-IR), X-ray diffractometry (XRD), and scanning electron microscopy (SEM) to analyze the resulting membranes, it was found that the ENPs and the chitosan-collagen composite membranes were highly compatible. Thermal properties (TG, DTG and DSC) were employed to evaluate the changes in the structural and physical properties which reached 384.94 °C, and a residual mass of 4.21 ± 0.13%. The ENPs loaded membranes exhibited marginally improved antioxidant and microbial growth inhibitory activities reaching 57.29 ± 2.77% and 10.33 ± 0.65 mm respectively. Additionally, the respective protease employed for the degradation studies was successfully expressed in Pichia pastoris (GS115) host cells, with functional pH and temperature optima between pH 4.0–5.0 and 60 °C respectively, and specific activity of 8408.9 ± 305.6 U/mg. It also exhibited enhanced specificity to its substrates upon collagen, chitosan, Bovine serum albumin (BSA), casein and casein sodium salt degradation with specific activities of 1477 ± 99.1, 1177 ± 103, and 1051.6 ± 60.7 U/mg respectively. Furthermore, the composite membranes showed excellent biodegradability by the 6th day in a natural environment treated with the aspartic protease from Aspergillus fumigatus (AfAP). Thus, the developed nanocomposite membranes from chitosan/collagen exhibited great potential and can well mimic the functions of a new generation of biodegradable and sustainable membranes. [ABSTRACT FROM AUTHOR]

N-glycosylation alters the properties of different enzymes in different ways. Rhizopus homothallicus was first described as an environmental isolate from desert soil in Guatemala. A new gene encoding glucanase RhGlu16B was identified in R. homothallicus. It had high specific activity (9673 U/mg) when barley glucan was used as a substrate, and β-glucan is hemicellulose that is abundant in nature. RhGlu16B has only one N-glycosylation site in its Ala55-Gly64 loop. It was found that N-glycosylation increased its Tm value and catalytic efficiency by 5.1 °C and 59%, respectively. Adding N-glycosylation to the same region of GH16 family glucanases TlGlu16A (from Talaromyces leycettanus) increased its thermostability and catalytic efficiency by 6.4 °C and 38%, respectively. In a verification experiment using GH16 family glucanases BisGlu16B (from Bisporus) in which N-glycosylation was removed, N-glycosylation also appeared to promote thermostability and catalytic efficiency. N-glycosylation reduced the overall root mean square deviation of the enzyme structure, creating rigidity and increasing overall thermostability. This study provided a reference for the molecular modification of GH16 family glucanases and guided the utilization of β-glucan in hemicellulose. [ABSTRACT FROM AUTHOR]

To acquire a further utilization of mulberry seed, antioxidant peptides derived from defatted mulberry seed protein was prepared using a continue-flow microreactor with the synergistic effects of immobilized protease and nontoxic ionic liquid. The mulberry seed meal was pretreated with the combined simultaneous ultrasound treatment at a frequency combination of 20/28/40 kHz, solid–liquid ratio of 1:30, output power of 400 W, and temperature of 40 °C; the extraction rate of mulberry seed protein was up to 70.1%, 21.54% higher than that obtained with the single-frequency ultrasonic treatment (p < 0.05). The hydrolysis process of mulberry seed protein in the continue-flow microreactor was catalyzed by the immobilized protamex combined with choline chloride, the flow rate was 2.5 mL/min, the temperature was 50 °C, and the hydrolysis rate was up to 83.33%, which was 1.14 times higher than that without ionic liquid. Moreover, the immobilized protease can be used 12 times under continue-flow condition, and the enzyme activity was still more than 80% of the original enzyme. Four antioxidant peptides including AMDGK (896.24 Da), KFRAKDG (1109.47 Da), FPSNEPLLG (1287.15 Da), and SNVPNKEL (1005.14 Da) were obtained by ionic liquid-enhanced microfluidic hydrolysis with immobilized protease from mulberry seed protein, and AMDGK had not been reported before. These peptides exhibited strong scavenging activities on 2,2-diphenyl-1-picrylhydrazyl radical (DPPH˙) (EC50 were 2.64, 1.89, 2.16, and 1.34 mg/mL, respectively) and superoxide anion radicals (·O2−) (EC50 were 3.36, 2.78, 2.56, and 1.77 mg/mL, respectively). Thus, the defatted mulberry seed protein possesses good edible development potential and economic value in the production of antioxidant peptides. [ABSTRACT FROM AUTHOR]

Due to the scarcity of resources, alternatives to non-renewable resources have become more important. Chemical catalysts react violently and the conditions are harsh. Using biocatalysts at the water/oil (W/O) interface remains technically challenging and is rewarding. In this study, the Thermomyces lanuginosus lipase-poly coupling biocatalyst was prepared by the atom transfer radical polymerization "grafting-from" method. The TL–PNIPAAm conjugate was successfully prepared and circular dichroism showed a negligible change between the free enzyme and conjugated enzyme by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and transmission electron microscopy. Furthermore, the temperature and pH stability of the conjugate was better than that of free TL. The remaining activity of the TL–PNIPAAm conjugate for 10 h was approximately 20% higher than that of free TL after heat treatment (55 °C) and higher pH stability was obtained in the pH 4–10. The remaining activity of the TL–PNIPAAm conjugates was above 60% after recycling 7 times. In addition, the conversion of biodiesel from kitchen waste oil was 92.6% at 50 °C for 25 h with a conversion efficiency of 0.04 g/mL/h. Thus, a high stability and reusability biocatalyst and stabilizer at the interface of water-in-oil Pickering emulsions was successfully prepared. [ABSTRACT FROM AUTHOR]

A large number of defective mulberries are discarded each year because mulberries are easy to break. The red pigments from defective mulberries are recognized as the sustainable sources of anthocyanins extracted from nature. Cyanidin-3-O-rutinoside and cyanidin-3-O-glucoside are the main components of mulberry red pigments, accounting for 50% and 40% of the total, respectively. Cyanidin-3-O-glucoside exhibits anticancer, hypoglycemic, and liver and visceral protection properties. Cyanidin-3-O-glucoside can be prepared by enzymatically hydrolyzing the rhamnosidase bond of cyanidin-3-O-rutinoside. To obtain mulberry red pigment with a high purity of cyanidin-3-O-glucoside, immobilized α-L-rhamnosidase was added to the aqueous two-phase system to construct a liquid-liquid-solid three-phase enzyme catalytic system. After optimization, the three-phase system was composed of 27.12% (w/w) ethanol, 18.10% (w/w) ammonium sulfate, 15% (w/w) mulberry juice, 4.24% (w/w) immobilized α-L-rhamnosidase, and 35.54% (w/w) pure water. The three-phase system was employed to enrich and purify cyanidin-3-O-glucoside at pH 5 and 45 °C for 1 h. The purity of cyanidin-3-O-glucoside was increased from 40 to 82.42% with cyanidin-3-O-rutinoside conversion of 60.68%. The immobilized α-L-rhamnosidase could be reused seven times, maintaining a relative activity of over 50%. Overall, the developed system provided an efficient and simple approach for high purity mulberry red pigment production and recycling in the field of sustainable agriculture. [ABSTRACT FROM AUTHOR]

Silkworm pupae are a typical byproduct with high protein content from silk-reeling industry. The aim of this study was to evaluate the performance of defatted silkworm pupae hydrolysate as an alternative nitrogen source to produce polysaccharides and flavonoids using the fungus Phellinus baumii. The results indicated that the defatted silkworm pupa was treated with dispase, and the freeze-dried hydrolysate had a yield of 37% and a nitrogen content of 3.18%. In the fermentation process, the content of the hydrolysates was 32 g/L. Moreover, the consuming reducing sugar accounted for 75.61% of the total reducing sugar and the nitrogen consumption accounted for 35.19% of the total nitrogen content. The maximum biomass produced by the hydrolysate treatment group was 0.6 g/L, and the extracellular flavonoid concentration was 0.4 g/L; the produced extracellular polysaccharide concentration was 7.71 g/L, which was 1.4–5.9 times higher than that of other nitrogen source treatment groups. Therefore, the defatted silkworm pupae hydrolysate is an effective alternative nitrogen source which has great economic value. As a nitrogen source reagent, it enhanced the fermentation ability of P. baumii and increased the yield of extracellular polysaccharides and flavonoids. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum is a common plant bacterium, and the improper use of chemicals to control this bacterium is harmful to the environment. Many undeveloped aromatic plants and green plant waste can be used for the preparation of essential oils (EOs). EOs have various biological activities, such as bacteriostatic, insecticidal and anti-oxidation, and are currently in the exploration stage for the control of R. solanacearum. In the present study, the inhibitory activity of ten EOs against R. solanacearum was determined by an antibacterial test. To prolong the duration of antibacterial activity, microcapsules of EOs were prepared by the complex coacervation method, and the optimal preparation conditions were obtained by central composite design method. The results showed that cinnamon EOs had the strongest antibacterial activity. For GIM1.71, GIM1.74, GIM1.76, GMI1000 and RS-5, the EC50 values of cinnamon EOs were 0.055 mg/mL, 0.071 mg/mL, 0.03 mg/mL, 0.067 mg/mL and 0.063 mg/mL, respectively. The preparation condition of cinnamon EOs microcapsules was as follows: the core-wall ratio was 4.35:1, the ratio of chitosan and sodium alginate was 0.15:1, and the calcium chloride concentration was 0.95 wt%. The encapsulation efficiency was 87.32%, and the average particle size of microcapsule was 870.4 μm. The release rate of microcapsule was 44.53% at 100 °C for 6 h and the release of the core materials in microcapsules was in line with Korsmeyer–Peppas release kinetics. Microcapsules placed at room temperature for 7 days still had significant antibacterial activity against five species of R. solanacearum. Due to its good stability and antibacterial activity, cinnamon EOs microcapsules will have great market application prospects as antibacterial agents. [ABSTRACT FROM AUTHOR]

Lipids produced from agricultural and industrial residues using oleaginous microorganisms for use as biofuels are attracting the attention of researchers due to their environmental benefits. However, low efficiencies and high costs limit their application to a certain extent. The present study is the first to use inositol as an enhancer to improve the production and accumulation of lipids during fermentation by the microalga Schizochytrium sp. SR21. The study aimed to maximize the production of lipids and docosahexaenoic acid (DHA) by optimizing the conditions of inositol addition into the fermentation medium. The corresponding key enzyme and metabolite profiles of SR21 were evaluated. The results indicated that the addition of 120 mg L−1 of inositol to the medium at 48 h improved lipid and DHA production by 13.90 and 20.82%, resulting in total concentrations of 22.86 and 8.53 g/L, respectively. Moreover, the ratio of unsaturated fatty acids (UFAs) to saturated fatty acids (SFAs) increased by 23.38% and is consistent with the results of the metabolomic analysis. The activity of enzymes (i.e., PC, G6PDH, NADPH-ME, and ACL) related to fatty acid synthesis in strain SR21 also increased significantly (43.38%, 28.68%, 37.47%, and 19.87%, respectively). Metabolomic analysis also showed that inositol promoted lipid synthesis in SR21 and significantly increased the relative proportion of UFAs by affecting the citrate cycle and SFA and UFA metabolic pathways. Thus, inositol is an ideal enhancer of lipid production and accumulation by oleaginous microorganisms. [ABSTRACT FROM AUTHOR]