Pulsed Light Mutagenesis of the Willow Bracket Mushroom, Phellinus igniarius (Agaricomycetes), for Enhanced Production of Flavonoids, Laccase, and Fermentation Biomass.

Phellinus igniarius is a medicinal fungus possessing potent therapeutic activity due to the polysaccharides, polyphenols, flavonoids, and other secondary metabolites they contain. Laccases are crucial enzymes involved in lignin degradation in Ph. igniarius and offer great potential to accomplish several bioprocesses. To generate Ph. igniarius strains with high biomass, flavonoid, and laccase activity, we used pulsed light (PL) technology for mutagenesis of Ph. igniarius protoplasts and screened for mutants with high biomass, flavonoid, and laccase activity. At the irradiation power of 100 J, treated distance 8.5 cm, irradiation frequency was 0.5 s/time, three times treatments, after five generations of selection, three mutants were obtained with higher biomass production. Compared with control, the mycelium biomass and the flavonoid production of the screened mutant strain QB72 were increased 20.87% and 53.51%, respectively. The total amount of the accumulated extracellular laccase of the QB72 in the first 6 and 8 days increased 23.38% and 22.37% respectively, and over the total 16 days it increased 9.62%. In addition, RAPD analysis results indicated that the genetic materials of the mutant QB72 were altered. PL mutagenesis method has great potential for developing strains, especially Phellinus.

Harnessing the Keratinolytic Activity of Bacillus licheniformis Through Random Mutagenesis Using Ultraviolet and Laser Irradiations.

Keratinase is one of the important proteases, which is widely used for converting keratin of the keratinaceous materials into various value-added products. In this study, a popular keratinase producer, Bacillus licheniformis PWD-1, was exposed to ultraviolet (UV) and He-Ne laser irradiations to develop high keratinase-producing mutants. Laser irradiation showed a higher lethality of cells (94%) than UV treatment (92%), whereas laser treatment required a longer time (75 min) than UV treatment (20 min). A total of 58 mutants were selected from 176 isolates to study protein and keratinase production capability of the mutants. The highest keratin-to-casein (K:C) ratio (1.43) was exhibited by LU11 mutant, which was obtained from the combined laser and UV irradiations. The purified keratinase (65 kDa) of LU11 showed 40% yield 1.7-fold purity, while the respective value for wild enzyme was 29% and 1.3-fold. Both enzymes showed optimal activity at 55 ℃ and pH 8, with a Z value of 15.78 ℃ for LU11 and 19.72 ℃ for wild strain. The Vmax and specific constant (Kcat/Km) of the mutant enzyme were 357.17 U/ml and 33.11 min-1 mM-1, respectively, which were significantly higher than the respective values of wild enzyme (102.04 U/ml and 28.36 min-1 mM-1).

Enhanced Mycelium Production of Phellinus igniarius (Agaricomycetes) Using a He-Ne Laser with Pulsed Light.

This study used a He-Ne laser with pulsed light irradiation to produce mutant strains of Phellinus igniarius strain JQ9 with enhanced characteristics for fermentation (17.685 ± 3.092 g/L) compared with the parent strain (12.062 ± 1.119 g/L). The combined treatment conditions were as follows: He-Ne laser irradiation for 30 min using a spot diameter of 10 mm, pulsed light treatment power set at 100 J, a treatment distance of 14.5 cm, and a flash frequency of 0.5 s. The production of bioactive polysaccharides and small biocompounds such as polyphenols, flavonoids, and triterpenes increased together with mycelium production. The results showed that polyphenol content was significantly correlated with L*, a*, and b* values (R = -0.594, P < 0.01; R = 0.571, P < 0.01; and R = 0.500, P < 0.05; respectively). Antagonistic and random amplified polymorphic DNA analyses indicated that the genetic material of the screened mutants was altered. The mutant screening using a He-Ne laser with pulsed light irradiation could be an effective method for the development of Phellinus strains and could thus improve mycelium production.

Effect of multi-frequency countercurrent ultrasound treatment on extraction optimization, functional and structural properties of protein isolates from Walnut (Juglans regia L.) meal.

This study evaluated the effects of ultrasound treatment on walnut meal protein (WMP) extraction and techno-functional properties. The Box-Behnken Design (BBD) was adopted for the optimization of the traditional and ultrasound-assisted extraction (UAE) processes. Standard protocols were used to assay the techno-functional characteristics. The extraction models' statistical results exhibited adequacy with the least desirability index of 95.8%. The UAE enhanced the WMP extraction yield, purity, and chemical score by 30.15%, 16.27%, and 9.74%, respectively, while reducing the extraction time by 25% over the control. The emulsion and foam stabilities and bulk density increased by 34.5%, 39.8%, and 6.1%, respectively, over the control. The α-helix decreased while ß-sheet, ß-turns and random coil secondary structure components increased significantly (p < .05) by 95.76%, 101.3%, 105.1%, and 85.7% correspondingly. The dual-frequency combination (20/40 kHz/kHz) was the best frequency mode. WMP could serve as a functional additive in manufactured foods as texture and flavor enhancer. PRACTICAL APPLICATIONS: Walnut meal protein (WMP) has a well-balanced amino acid profile and its economic use could be practically increased as a food ingredient by ultrasound-assisted extraction. By this technique, WMP could be employed for the development of enhanced food ingredients rather than being discarded as animal feed. This study showed a positive effect of ultrasonic-assisted alkaline pretreatment on WMP extraction, functionality and structure characteristics. In addition to process improvement, ultrasound is energy efficient and environmentally friendly. Therefore, the applicability of this technique to improve the functionality of plant proteins from industrial by-products to be included in food products is promising.

Shelf-life extension of grape (Pinot noir) by xanthan gum enriched with ascorbic and citric acid during cold temperature storage.

The detrimental health implications of chemical preservatives in fruits have necessitated exploitation of safe and natural alternatives such as edible gums. This work studied shelf-life extension in grape (Pinot noir) under cold storage by xanthan gum (XAN) coatings enriched with ascorbic acid (XANAS) and citric acid (XANCI). Standard scientific methods were used to examine some sensory (color, texture-resilience and hardness), enzyme, anthocyanine and antioxidant activities. Also, the reaction rate mechanism was examined through modeling of selected shelf-life indicators; color change, weight loss, and antioxidants. The results revealed that, Xanthan gum and its acid modified coatings significantly (p < 0.05) suppressed polyphenol oxidase, ascorbic acid oxidase, polymethyl etherase acitivies and maintained the structural integrity of the grape during the 21 days storage period. Weight loss (%) in the grape samples was 13.66 < 13.98 < 14.16 < 15.64 in the order XANAS < XANCI < XAN < CONTROL whilst ferric reducing antioxidant power (FRAP) activity was 150.23 > 143.18 > 136.49 > 104.5 mg/100 g AEAC corresponding to XAN > XANAS > XANCI > CONTROL. Significantly (p < 0.05) higher phytochemical contents were observed in the gum coatings compared to the control. Through statistical parameters such as the coefficient of determination (R 2), root mean square error (RMSE) and reduced Chi square (χ2), the second-order polynomial model predicted precisely the decomposition of color, weight loss and FRAP of grape. Color deterioration was attributed to changes in b* parameter as a result of phenolics and phytochemical decompositions resulting from enzymatic activities. Conclusively, acid modified xanthan gum coatings could preserve phytochemicals, color, antioxidant and textural properties of grape in cold temperature storage.

Sonochemical action and reaction of edible insect protein: Influence on enzymolysis reaction-kinetics, free-Gibbs, structure, and antioxidant capacity.

We investigated the impact of sonochemical action and the reaction of Hermetia illucens larvae meal protein (HILMP) as regards enzymolysis under varied enzyme concentration and temperature to explain the mechanism and effect of sonication on molecular conformation, limits of kinetics, free-Gibbs energy, and antioxidative capacity. Control treatment was used for comparison. The results showed sonochemical treatment enhanced HILMP-enzymolysis efficiency at various enzyme volume, and temperature. Enzymolysis-kinetics revealed sonochemical treatment increased the rate constant (p < .05) by 17.21%, 25.06%, 26.91%, and 41.38% at 323, 313, 303, and 293 K, respectively. On free-Gibbs, sonochemical treatment reduced the reactants-reactivity energy, enthalpy, and entropy by 30.53%, 35.05%, and 10.71%, respectively (p < .05). Changes in spectra of UV and fluorescence, and micrographic imaging indicated alterations of HILMP by sonochemical treatment. Antioxidative activity of sonochemically-treated HILMP increased, compared to control. Thus, sonochemical treatment may be beneficial in the production of edible insect proteins with smaller molecular weights for different food and/or pharmaceutical applications. PRACTICAL APPLICATIONS: Sonochemical pretreatment of HILMP positively impacted it enzymolysis rate-reaction, stability of reaction products, structure, and bioactivity. Thus, the technique may be beneficial to industry in the processing/development of new (bioactive/pharmaceutical) products involving enzymolysis of edible insects (e.g., Hermetia illucens) protein; particularly at such a time where edible insects are projected to be a source of protein for human nutrition and livestock in the next few years.

Enzymolysis of walnut (Juglans regia L.) meal protein: Ultrasonication-assisted alkaline pretreatment impact on kinetics and thermodynamics.

To improve the utility of walnut meal protein, influences of ultrasonication on the kinetic and thermodynamic constraints of enzyme (trypsin) hydrolysis were assessed using a multi-frequency counter current S-type ultrasound machine. Results revealed that both the ultrasonication treatment and control hydrolysis obeyed the first-order kinetics within the study constraints. After 90 min hydrolysis time, the uppermost percentage conversion rate (57.5%) of substrate to product and the percentage increase (25.42%) in hydrolyzed protein concentration were obtained at 323 K and 10% substrate concentration, respectively. The Michaelis constant (KM ) in ultrasonic enzymolysis declined by 58.66% over the control. The reaction rate coefficient (k) for the proteolysis improved by 84.75%, 52.43%, 48.25%, and 37.79% at 219, 303, 313, and 323 K, correspondingly. Generally, the bond energy (Ea ), enthalpy change (ΔH), entropy change (ΔS), and Gibbs free energy (ΔG) were reduced by the ultrasound pretreatment with 36.61%, 28.05%, 18.22%, and 5.24%, respectively. PRACTICAL APPLICATIONS: Walnut meal protein has a well-balanced amino acid profile and its economic utilization could be increased practicably as a food ingredient via production of hydrolysates/peptides for the production of improved food ingredients instead of being cast off as animal feed. This study demonstrated a positive bearing of ultrasonication-assisted alkaline pretreatment on proteolytic reaction characteristics and its energy efficiency for walnut meal protein, which makes this technique applicable to the enhancement of plant proteins for inclusion in food products, especially tree nut and oil seed waste products from the oil industry.

Modeling of drying and ameliorative effects of relative humidity (RH) against ß-carotene degradation and color of carrot (Daucus carota var.) slices.

Drying and ß-carotenes retention kinetics were predicted using models in relative humidity (RH) drying condition. This was achieved by drying carrot slices using RH-convective hot-air dryer at 60, 70 and 80 °C under RH (10% 20% and 30%) conditions at 2.0 m/s air velocity. Three mathematical models describing thin layer were compared to their goodness of fit in terms of coefficient of correlation (R2), root mean square error (RMSE) and reduced Chi square ( χ 2 ). The Wang and Singh model could satisfactorily describe RH-convective drying of carrot slices with R2, RMSE and χ 2 in the ranges of 0.996-0.999, 5.4 × 10-4-9.4 × 10-4 and 0.0150-0.03353 respectively. The results reveal that a range of 3.61-8.2% retention of ß-carotene was observed for every 10% increase in RH in various drying air temperature. In summary, higher temperatures were mainly responsible for ß-carotenes degradation however this can be mitigated when drying is conducted under higher RH.

Feasibility study on direct fermentation of soybean meal by Bacillus stearothermophilus under non-sterile conditions.

BACKGROUND: To evaluate the feasibility of high-temperature solid-state fermentation (SSF) using soybean meal (SBM) during the non-sterile process, Bacillus stearothermophilus was employed to assess the nutritional quality and bioactivity of SBM after fermentation. RESULTS: The fermented SBM (FSBM) without autoclaving showed significant improvements in nutritional quality and bioactivity. The contents of peptides and crude and soluble proteins increased by 131.21%, 5.3% and 15.52%, respectively. Meanwhile, DPPH (2,2-diphenyl-1-picrylhydrazyl) scavenging ability, reducing ability and hydroxyl free radical-scavenging activity rose by 57.07%, 238.92% and 368.26%, respectively. The inhibitory activity of angiotensin I-converting enzyme increased from 1.43 ± 0.83% to 26.89 ± 1.03%, while the trypsin inhibitor activity decreased by 74.05%. The contents of neutral and alkaline proteases and the growth of microorganisms in FSBM without autoclaving were higher and better than in steam-treated FSBM. After steam treatment, the water-holding capacity of SBM decreased, and a high crosslink density was observed on the surface of SBM particles. CONCLUSIONS: It is feasible to ferment SBM by high-temperature SSF using B. stearothermophilus under non-sterile conditions. Adverse effects of SSF using sterile SBM might be owing to the low water-holding capacity caused by autoclaving. © 2018 Society of Chemical Industry.