Enzyme Production by Solid Substrate Fermentation of <Emphasis Type="Italic">Pleurotus ostreatus</Emphasis> and <Emphasis Type="Italic">Trametes versicolor</Emphasis> on Tomato Pomace

A process of solid state fermentation (SSF) on tomato pomace was developed with the white-rot fungi Pleurotus ostreatus and Trametes versicolor, using sorghum stalks as support. Operative parameters (humidity, water activity, and size of substrate particles) guaranteeing a good colonization of tomato pomace by both fungi were defined and conditions for production at high titers of the industrially relevant enzymes laccase, xylanase and protease were identified. Significant laccase activity levels (up to 36 U g⁻¹ dry matter) were achieved without any optimization of culture conditions, neither by nutrient addition nor by O₂ enrichment. Furthermore, protease activity levels up to 34,000 U g⁻¹ dry matter were achieved, being higher than those reported for the fungi typically considered as the best protease producers such as Aspergillus strains. Moreover, as one of the most significant results of this study, analysis of P. ostreatus tomato SSF samples by zymogram revealed two bands with laccase activity which had not been detected so far.     

Production of lipase by a newly isolated <Emphasis Type="Italic">Bacillus coagulans</Emphasis> under solid-state fermentation using melon wastes

Summary An extracellular lipase was produced by Bacillus coagulans by solid-state fermentation. Solid waste from melon was used as the basic nutrient source and was supplemented with olive oil. The highest lipase production (78,069 U/g) was achieved after 24h of cultivation with 1% olive oil enrichment. Enzyme had an optimal activity at 37°C and pH 7.0, and sodium dodecyl sulfate increased lipase activity. NH ₄NO₃ increased enzyme production, whereas organic nitrogen had no effect. The effect of the type of carbon sources on lipolytic enzyme production was also studied. The best results were obtained with starch and maltose (148,932 and 141,629 U/g, respectively), whereas a rather low enzyme activity was found in cultures grown on glucose and galactose (approx 118,769 and 123,622 U/g, respectively). Enzyme was inhibited with Mn⁺² and Ni⁺² by 68 and 74%, respectively. By contrast, Ca⁺² enhanced enzyme production by 5%.     

Production and Characterization of an Alkaline Thermostable Crude Lipase from an Isolated Strain of <Emphasis Type="Italic">Bacillus cereus</Emphasis> C<Subscript>7</Subscript>

A bacterial strain isolated from spoiled coconut and identified as Bacillus cereus was found capable of producing alkaline thermostable extracellular lipase. Optimum temperature, time, and pH for enzyme substrate reaction were found to be 60 °C, 10 min, and 8.0 respectively. Common surfactants except Triton X 100 and cetyltrimethylammonium bromide have no or very little inhibitory effects on enzyme activity. The enzyme was found to be stable in presence of oxidizing agents and protease enzyme. The maximum lipase production was achieved at 30–33 °C, pH 8.0 on 24 h of fermentation using 50 ml medium in a 250-ml Erlenmeyer flask. The superior carbon and nitrogen sources for lipase production were starch (2%) and ammonium sulfate (nitrogen level 21.2 mg/100 ml), peptone (nitrogen level 297 mg/100 ml), and urea (nitrogen level 46.62 mg/100 ml) in combination, respectively. The maximum enzyme activity obtained was 33 ± 0.567 IU/ml.     

Keratinase Production by Endophytic <Emphasis Type="Italic">Penicillium</Emphasis> spp. Morsy1 Under Solid-State Fermentation Using Rice Straw

Among all endophytic keratinolytic fungal isolates recovered from marine soft coral Dendronephthya hemprichii, Penicillium spp. Morsy1 was selected as the hyperactive keratinolytic strain under solid substrate fermentation of different agriculture and poultry wastes. The optimization of extraction process, physicochemical parameters affecting the keratinase production in solid-state fermentation, and the purified keratinase parameters were studied. Maximum keratinase activity (1,600 U g⁻¹, initial dry substrate) was recovered from moldy bran with 0.1% Tween 80. The optimized production conditions were rice straw as carbon source, pH of medium 6, growth temperature 26 °C, initial moisture content of 80% (v/w), inoculum size of 10⁵ spores ml⁻¹, and an average particle size of the substrate 0.6 mm (3,560 U g⁻¹, initial dry substrate after 5 days of fermentation). Two types of keratinase (Ahm1 and Ahm2) were purified from the culture supernatant through ammonium sulfate precipitation, DEAE-Sepharose, and gel filtration chromatography. Enzyme molecular weights were 19 kDa (Ahm1) and 40 kDa (Ahm2). The kinetic parameters of purified keratinases were optimized for the hydrolysis of azokeratin by Ahm1 (pH 7.0–8.0, stable in pH range of 6.0 to 8.0 at 50 °C) and Ahm2 enzymes (pH 10.0–11.0, stable in pH range of 6.0 to 11.0 at 60–65 °C). Whereas inhibitors of serine (phenylmethylsulfonyl fluoride) and cysteine (iodoacetamide) proteases had minor effects on both Ahm1 and Ahm2 activity, both keratinases were strongly inhibited by chelating agents EDTA and EGTA. These findings suggest that serine and cysteine residues are not involved in the catalytic mechanisms, and they are metalloproteases.     

Xylanase Production by <Emphasis Type="Italic">Penicillium canescens</Emphasis> on Soya Oil Cake in Solid-State Fermentation

There is an increasing interest for the organic residues from various sectors of agriculture and industries over the past few decades. Their application in the field of fermentation technology has resulted in the production of bulk chemicals and value-added products such as amino acid, enzymes, mushroom, organic acids, single-cell protein, biologically active secondary metabolites, etc. (Ramachandran et al., Bioresource Technology 98:2000–2009, 2007). In this work, the production of extracellular xylanase by the fungus Penicillium canescens was investigated in solid-state fermentation using five agro-industrial substrates (soya oil cake, soya meal, wheat bran, whole wheat bran, and pulp beet). The best substrate was the soya oil cake. In order to optimize the production, the most effective cultivation conditions were investigated in Erlenmeyer flasks and in plastic bags with 5 and 100 g of soya oil cake, respectively. The initial moisture content, initial pH, and temperature of the culture affected the xylanase synthesis. The optimal fermentation medium was composed by soya oil cake crushed to 5 mm supplemented with 3% and 4% (w/w) of casein peptone and Na₂HPO₄.2H₂O. After 7 days of incubation at 30 °C and under 80% of initial moisture, a xylanase production level of 18,895 ± 778 U/g (Erlenmeyer flasks) and 9,300 ± 589 U/g (plastic bags) was reached. The partially purified enzyme recovered by ammonium sulfate fractionation was completely stable at freezing and refrigeration temperatures up to 6 months and reasonably stable at room temperature for more than 3 months.     

Banana Peel: A Potential Substrate for Laccase Production by <Emphasis Type="Italic">Aspergillus fumigatus</Emphasis> VkJ2.4.5 in Solid-State Fermentation

In solid-state fermentation, among various solid supports evaluated, banana peel was found to be an ideal support and resulted into higher levels of laccase (6281.4 ± 63.60 U l⁻¹) along with notable levels of manganese peroxidase production (1339.0 ± 131.23 U l⁻¹) by Aspergillus fumigatus VkJ2.4.5. Maximum levels of laccase was achieved under derived conditions consisting of 80% of moisture level, 6 days of incubation period, 6% inoculum level, and an aeration level of 2.5 l min⁻¹. A column-tray bioreactor was designed to scale up and economize the enzyme production in three successive cycles of fermentation using the same fungal biomass. Thermal and pH stability profiles revealed that enzyme was stable up to 50°C and at varying pH range from 5–9 for up to 2 h. The apparent molecular weight of laccase was found to be 34 ± 1 kDa. MALDI-TOF/TOF analysis of the protein showed significant homology with maximum identity of 67% to other laccases reported in database.     

Cellulases and Xylanases Production by <Emphasis Type="Italic">Penicillium echinulatum</Emphasis> Grown on Sugar Cane Bagasse in Solid-State Fermentation

To investigate the production of cellulases and xylanases from Penicillium echinulatum 9A02S1, solid-state fermentation (SSF) was performed by using different ratios of sugar cane bagasse (SCB) and wheat bran (WB). The greatest filter paper activity obtained was 45.82 ± 1.88 U gdm⁻¹ in a culture containing 6SCB/4WB on the third day. The greatest β-glucosidase activities were 40.13 ± 5.10 U gdm⁻¹ obtained on the third day for the 0SCB/10WB culture and 29.17 ± 1.06 U gdm⁻¹ for the 2SCB/8WB culture. For endoglucanase, the greatest activities were 290.47 ± 43.57 and 276.84 ± 15.47 U gdm⁻¹, for the culture 6SCB/4WB on the fourth and fifth days of cultivation, respectively. The greatest xylanase activities were found on the third day for the cultures 6SCB/4WB (36.38 ± 5.38 U gdm⁻¹) and 4SCB/6WB (37.87 ± 2.26 U gdm⁻¹). In conclusion, the results presented in this article showed that it was possible to obtain large amounts of cellulases and xylanases enzymes using low-cost substrates, such as SCB and WB.     

Significant Improvement of <Emphasis Type="Italic">Serratia marcescens</Emphasis> Lipase Fermentation,by Optimizing Medium,Induction, and Oxygen Supply

Production of an extracellular lipase from Serratia marcescens ECU1010, which is an industrially important biocatalyst for the stereospecific synthesis of Diltiazem precusor, was carefully optimized in both shake flasks and a fermenter, using Tween-80 as the enzyme inducer. Dextrin and beef extract combined with ammonium sulfate were indicated to be the best carbon and nitrogen sources, respectively. With the increase of Tween-80 from 0 to 10 g l⁻¹, the lipase production was greatly enhanced from merely 250 U l⁻¹ to a maximum of 3,340 U l⁻¹, giving the highest lipase yield of ca 640 U g⁻¹ dry cell mass (DCW), although the maximum biomass (6.0 g DCW l⁻¹) was achieved at 15 g l⁻¹ of Tween-80. When the medium loading in shake flasks was reduced from 20 to 10% (v / v), the lipase production was significantly enhanced. The increase in shaking speed also resulted in an improvement of the lipase production, although the cell growth was slightly repressed, suggesting that the increase of dissolved oxygen (DO) concentration contributed to the enhancements of lipase yield. When the lipase fermentation was carried out in a 5-l fermenter, the lipase production reached a new maximum of 11,060 U l⁻¹ by simply raising the aeration rate from 0.5 to 1.0 vvm, while keeping the dissolved oxygen above 20% saturation via intermittent adjustment of the agitation speed (≥400 rpm), in the presence of a relatively low concentration (2 g l⁻¹) of Tween-80 to prevent a potential foaming problem, which is easy to occur in the intensively aerated fermenter.     

Valorization of By-Products from Palm Oil Mills for the Production of Generic Fermentation Media for Microbial Oil Synthesis

This study demonstrates the production of a generic nutrient-rich feedstock using by-product streams from palm oil production that could be used as a substitute for commercial fermentation supplements. Solid-state fermentations of palm kernel cake (PKC) and palm-pressed fiber (PPF) were conducted in tray bioreactors and a rotating drum bioreactor by the fungal strain Aspergillus oryzae for the production of crude enzymes. The production of protease was optimized (319.3 U/g) at an initial moisture content of 55 %, when PKC was used as the sole substrate. The highest free amino nitrogen (FAN) production (5.6 mg/g) obtained via PKC hydrolysis using the crude enzymes produced via solid-state fermentation was achieved at 50 °C. Three initial PKC concentrations (48.7, 73.7, and 98.7 g/L) were tested in hydrolysis experiments, leading to total Kjeldahl nitrogen to FAN conversion yields up to 27.9 %. Sequential solid-state fermentation followed by hydrolysis was carried out in the same rotating drum bioreactor, leading to the production of 136.7 U/g of protease activity during fermentation and 196.5 mg/L of FAN during hydrolysis. Microbial oil production was successfully achieved with the oleaginous yeast strain Lipomyces starkeyi DSM 70296 cultivated on the produced PKC hydrolysate mixed with commercial carbon sources, including glucose, xylose, mannose, galactose, and arabinose.     

High-yield bacillus subtilis protease production by solid-state fermentation

A Bacillus subtilis isolate was shown to be able to produce extracellular protease in solid-state fermentations (SSF) using soy cake as culture medium. A significant effect of inoculum concentration and physiological age on protease production was observed. Maximum activities were obtained for inocula consisting of exponentially growing cells at inoculum concentrations in the range of 0.7–2.0 mg g⁻¹. A comparative study on the influence of cultivation temperature and initial medium pH on protease production in SSF and in submerged fermentation (SF) revealed that in SSF a broader pH range (5–10), but the same optimum temperature (37°C), is obtained when compared to SF. A kinetic study showed that enzyme production is associated with bacterial growth and that enzyme inactivation begins before biomass reaches a maximum level for both SF and SSF. Maximum protease activity and productivity were 960 U g⁻¹ and 15.4 U g⁻¹ h⁻¹ for SSF, and 12 U mL⁻¹ and 1.3 U mL⁻¹ h⁻¹ for SF. When SSF protease activity was expressed by volume of enzyme extract, the enzyme level was 10-fold higher and the enzyme productivity 45% higher than in SF. These results indicate that this bacterial strain shows a high biotechnological potential for protease production in solid-state fermentation.     

Silk-Fiber Immobilized Lipase-Catalyzed Hydrolysis of Emulsified Sunflower Oil

Lipase was immobilized in silk fibers through glutaraldehyde cross-linking to a maximum loading of 59 U/g silk-fiber and the immobilized lipase was utilized for the hydrolysis of sunflower oil (Helianthus annuus). The hydrolytic activity of the lipase, which was poor in biphasic oil in water system, was increased significantly when the sunflower oil was emulsified in aqueous medium. The hydrolytic activities of the immobilized lipase were 48.73 ± 1.26 U, 36.11 ± 0.96 U, and nil when the substrate sunflower oil was used as emulsion created by a rhamnolipid biosurfactant, Triton X100, and ultrasonication, respectively. Although the efficiency of the immobilized lipase was less than 12% than the corresponding free lipase, the immobilized lipase could be reused for the biosurfactant-mediated hydrolysis of sunflower oil up to third cycle of the reaction. The yield of the fatty acids in the second, third, and fourth cycles were 49.45%, 22.91%, and 5.09%, respectively, of the yield obtained in the first cycle.     

Production of Lipids Containing High Levels of Docosahexaenoic Acid by a Newly Isolated Microalga, <Emphasis Type="Italic">Aurantiochytrium</Emphasis> sp. KRS101

In the present study, a novel oleaginous Thraustochytrid containing a high content of docosahexaenoic acid (DHA) was isolated from a mangrove ecosystem in Malaysia. The strain identified as an Aurantiochytrium sp. by 18S rRNA sequencing and named KRS101 used various carbon and nitrogen sources, indicating metabolic versatility. Optimal culture conditions, thus maximizing cell growth, and high levels of lipid and DHA production, were attained using glucose (60 g l⁻¹) as carbon source, corn steep solid (10 g l⁻¹) as nitrogen source, and sea salt (15 g l⁻¹). The highest biomass, lipid, and DHA production of KRS101 upon fed-batch fermentation were 50.2 g l⁻¹ (16.7 g l⁻¹ day⁻¹), 21.8 g l⁻¹ (44% DCW), and 8.8 g l⁻¹ (40% TFA), respectively. Similar values were obtained when a cheap substrate like molasses, rather than glucose, was used as the carbon source (DCW of 52.44 g l⁻¹, lipid and DHA levels of 20.2 and 8.83 g l⁻¹, respectively), indicating that production of microbial oils containing high levels of DHA can be produced economically when the novel strain is used.     

Production of Thermophilic Endo-β-1,4-xylanases by <Emphasis Type="Italic">Aspergillus fumigatus</Emphasis> FBSPE-05 Using Agro-industrial By-products

In the present paper, endo-β-1,4-xylanase production by Aspergillus fumigatus was evaluated in solid-state fermentation using low-cost substrates such as sugarcane bagasse (SCB), brewer’s spent grain (BSG), and wheat bran (WB). The partial characterization of the crude enzyme was also performed. In the experimental conditions, the highest levels of endo-β-1,4-xylanase production by A. fumigatus FBSPE-05 occurred within 8 days incubation when using SCB/liquid medium at 1:2 ratio (219.5 U g⁻¹) and 4 days incubation when using WB/liquid medium at 1:1 ratio (215.6 U g⁻¹). Crude enzyme from this last condition was used to enzyme characterization, showing best enzyme activity at 60 °C and pH 6.0, which suggests a thermophilic endoxylanase. The crude enzyme retained 73% of its activity after 1 h at 60 °C, and zymogram has shown three bands of endo-β-1,4-xylanase activity, with different molecular masses. A. fumigatus FBSPE-05 was able to grow and produce good levels of endo-β-1,4-xylanase using agro-industrial by-products, making this strain worthy for further investigation. To our knowledge, this is the first study reporting the use of SCB and/or BSG as sole substrates for endoxylanase production by solid-state fermentation using A. fumigatus.     

Integrated Use of Residues from Olive Mill and Winery for Lipase Production by Solid State Fermentation with Aspergillus sp.

Two-phase olive mill waste (TPOMW) is presently the major waste produced by the olive mill industry. This waste has potential to be used as substrate for solid state fermentation (SSF) despite of its high concentration of phenolic compounds and low nitrogen content. In this work, it is demonstrated that mixtures of TPOMW with winery wastes support the production of lipase by Aspergillus spp. By agar plate screening, Aspergillus niger MUM 03.58, Aspergillus ibericus MUM 03.49, and Aspergillus uvarum MUM 08.01 were chosen for lipase production by SSF. Plackett–Burman experimental design was employed to evaluate the effect of substrate composition and time on lipase production. The highest amounts of lipase were produced by A. ibericus on a mixture of TPOMW, urea, and exhausted grape mark (EGM). Urea was found to be the most influent factor for the lipase production. Further optimization of lipase production by A. ibericus using a full factorial design (3²) conducted to optimal conditions of substrate composition (0.073 g urea/g and 25 % of EGM) achieve 18.67 U/g of lipolytic activity.     

A stable lipase fromCandida lipolytica

Although Upases have been intensively studied, some aspects of enzyme production like substrate uptake, catabolite repression, and enzyme stability under long storage periods are seldom discussed in the literature. This work deals with the production of lipase by a new selected strain ofCandida lipolytica. Concerning nutrition, it was observed that inorganic nitrogen sources were not as effective as peptone, and that oleic acid or triacylglycerides (TAG) were essential carbon sources. Repression by glucose and stimulation by oleic acid and long chain TAG (triolein and olive oil) were observed. Extracellular lipase activity was only observed at high levels at late stationary phase, whereas intracellular lipase levels were constant and almost undetectable during the cultivation period, suggesting that the produced enzyme was attached to the cell wall, mainly at the beginning of cultivation. The crude lipase produced by this yeast strain shows the following optima conditions: pH 8.0–10.0, temperature of 55°C. Moreover, this preparation maintains its full activity for at least 370 d at 5°C.     

Lipase from a Brazilian StrainPenicillium citrinum Cultured in a Simple and Inexpensive Medium st]Heat-Denaturation, Kinetics, and pH Stability

This work is a study of lipase production by a Brazilian strain ofPenicillium citrinum using an inexpensive and simple medium without organic nitrogen sources and of some important industrial properties, including thermostability in relation to ionic strength. The maximal lipase activity (1585 U/L) was obtained whenPenicillium citrinum was cultured on 0.75% ammonium sulfate complemented with minerals salts instead of yeast extract. Although this activity was about 55% lower than that produced in medium with yeast extract (2850 U/L), the specific activity (7.8 U/mg proteins) was higher than that obtained with the yeast extract (4.9 U/mg proteins). The morphology of fungus changed totally, with yeast extract there are smooth, solid, and spherical pellets whereas on ammonium sulfate there are small “hairy” pellets uniformly suspended in the medium. The effect of ferrous (Fe⁺⁺) ions was carried out using medium MA with and without Fe⁺⁺ ions. Lipase production byPenicillium citrinum in medium MA requires Fe⁺⁺ ions, the absence of which caused a decreased of about 50% in the specific activity (3.5 U/mg proteins). The utilization of commercial, locally available oils as carbon sources, such as soybean oil (236 U/L) and corn oil (74 U/L) resulted in lower activity compared to olive oil, showing that lipase production byPenicillium citrinum is specifically induced by olive oil. Potassium concentration in the medium can effects the production of lipase (1 mM (1585 U/L), 10 mM (1290 U/L), and 30 mM (1238 U/L), 50 mM (195 U/L), and 100 mM (2 U/L). The crude culture filtered was susceptable to thermal deactivation. It was stable at pH 6.0, but was not stable at the optimum pH (8.0-8.5) at 50 mM. At the low ionic concentration (1-25 mM) this lipase was stable at low pH (3.5-4.0). The activation energy was 22.4 ±2.2 Kcal. mol 1.     

Immobilization of Candida cylindracea Lipase by Covalent Attachment on Glu-Modified Bentonite

Alkaline Ca-bentonite, obtained upon acid activation and base load of natural bentonite, has a good anion exchange capability. Glu-modified alkaline Ca-bentonites were further prepared by covalent binding with glutamic acid for the immobilization of lipase OF from Candida cylindracea. The obtained immobilized lipase demonstrated a significantly higher catalytic activity than that of unmodified alkaline Ca-bentonite, giving a specific activity of 62.1 U mg⁻¹ protein, twice that of the unmodified carrier, and a total activity of 391.2 U g⁻¹ support, retaining ~ 82.3% of the activity after being reused five times for olive oil emulsion hydrolysis. X-ray diffraction and Fourier transform infrared spectroscopy assays demonstrated the successful immobilization of the lipase on the surface of the bentonite. Upon immobilization, the thermostability of the lipase improved remarkably. At 50 °C, free lipase retained only 6.0% of its initial activity at 6 h, in comparison with 15% for Ca-Bent-lipase and 50% for Glu-Ca-Bent-lipase after 8 h. The Glu-Ca-Bent-lipase is proved as an effective biocatalyst for the biodiesel preparation, improving the transesterification reaction conversion from 52.8% in the condition of free lipase to 99.9% and keeping at 56.2% after being reused five times, while the free lipase was inactive upon two reuses. The above results provide a new route in the use of inexpensive bentonite for the enzyme immobilization.

     

Scale up and Application of Biosurfactant from <Emphasis Type="Italic">Bacillus subtilis</Emphasis> in Enhanced Oil Recovery

There is a lack of fundamental knowledge about the scale up of biosurfactant production. In order to develop suitable technology of commercialization, carrying out tests in shake flasks and bioreactors was essential. A reactor with integrated foam collector was designed for biosurfactant production using Bacillus subtilis isolated from agricultural soil. The yield of biosurfactant on biomass (Y _p/x), biosurfactant on sucrose (Y _p/s), and the volumetric production rate (Y) for shake flask were obtained about 0.45 g g⁻¹, 0.18 g g⁻¹, and 0.03 g l⁻¹ h⁻¹, respectively. The best condition for bioreactor was 300 rpm and 1.5 vvm, giving Y _x/s, Y _p/x, Y _p/s, and Y of 0.42 g g⁻¹, 0.595 g g⁻¹, 0.25 g g⁻¹, and 0.057 g l⁻¹ h⁻¹, respectively. The biosurfactant maximum production, 2.5 g l⁻¹, was reached in 44 h of growth, which was 28% better than the shake flask. The obtained volumetric oxygen transfer coefficient (K _L a) values at optimum conditions in the shake flask and the bioreactor were found to be around 0.01 and 0.0117 s⁻¹, respectively. Comparison of K _L a values at optimum conditions shows that biosurfactant production scaling up from shake flask to bioreactor can be done with K _L a as scale up criterion very accurately. Nearly 8% of original oil in place was recovered using this biosurfactant after water flooding in the sand pack.     

A Novel Thermostable and Alkaline Protease Produced from Bacillus stearothermophilus Isolated from Olive Oil Mill Sols Suitable to Industrial Biotechnology

This study was conducted to identify a new alkaline and thermophilic protease (Ba.St.Pr) produced from Bacillus stearothermophilus isolated from olive oil mill sols and to evaluate its culture conditions, including temperature, pH, carbon and nitrogen sources, and incubation time. The optimum culture conditions for cell growth (10 g/L) and protease production (5050 U/mL) were as follows: temperature 55 °C, pH 10, inoculation density 8 × 10⁸ CFU/mL, and incubation time 24 h. The use of 3% yeast extract as the nitrogen sources and galactose (7.5 g/L) as the carbon sources enhanced both cell growth and protease production. Using reversed-phase analytical HPLC on C-8 column, the new protease was purified with a molecular mass of approximately 28 kDa. The N-terminal sequence of Ba.St.Pr exhibited a high level of identity of approximately 95% with those of Bacillus strains. Characterization under extreme conditions revealed a novel thermostable and alkaline protease with a half-life time of 187 min when incubated with combined Ca²⁺/mannitol. Ba.St.Pr demonstrated a higher stability in the presence of surfactant, solvent, and Ca²⁺ ions. Consequently, all the evaluated activity parameters highlighted the promising properties of this bacterium for industrial and biotechnological applications.     

High-Yield Hypocrellin A Production in Solid-State Fermentation by <Emphasis Type="Italic">Shiraia</Emphasis> sp. SUPER-H168

Hypocrellin A production by Shiraia sp. SUPER-H168 was studied under solid-state fermentation. Corn was found to be the best substrate after evaluating eight kinds of agro-industrial crops and residues. The optimized solid-state fermentation conditions were as follows: inoculum size 3 × 10⁶ spores, substrate particle size 0.8–1 mm, initial moisture content 50%, and temperature 30 °C. Six kinds of external carbon source and seven kinds of external nitrogen source were evaluated, respectively, for HA production. Glucose and NaNO₃ were the best. The combination of them was optimized by the response surface method. The optimum compositions of the supplementary glucose and NaNO₃ were 1.65 g/100 g and 0.43 g/L, respectively. Hypocrellin A production reached 4.7 mg/g.