Rhizopus Oryzae Research Articles

Antifungal Activity and Mechanism of Electron Beam Irradiation Against Rhizopus oryzae

Electron beam irradiation is a physical fungicidal technique that has emerged as a potential application in China. However, its antifungal activity and mechanism against Rhizopus oryzae have not been reported. Thus, this study aimed to investigate the antifungal activity and mechanism of electron beam irradiation of R. oryzae. The antifungal activity analysis showed that the D10 value and complete elimination dose of R. oryzae irradiated by electron beam were 1.73 kGy and 8.08 kGy, respectively. Electron beam irradiation has a strong inhibitory effect on the filamentous biomass of R. oryzae. To reveal the antifungal mechanism of electron beam against R. oryzae, this study analyzed the dynamic changes in the cell wall, cell membrane, and oxidative stress induced by different irradiation doses. The results showed that electron beam irradiation destroyed the cell wall structure of R. oryzae, increasing chitinase activity and decreasing chitin content. Cell membrane integrity is disrupted, increasing relative conductivity, decreasing pH values, and decreasing soluble protein content. Electron beam irradiation causes oxidative stress in cells, increasing H2O2 content, decreasing antisuperoxide anion activity, decreasing DPPH free radical scavenging activity, and inhibiting defense enzyme (CAT and SOD) activity. This phenomenon indicates that electron beams can cause structural damage to and metabolic dysfunction of cells and disorders of redox homeostasis, which may be the main cause of growth inhibition and cell death in R. oryzae.

Investigating the induction of polyphenol biosynthesis in the cultured Cycolocarya paliurus cells and the stimulatory mechanism of co-induction with 5-aminolevulinic acid and salicylic acid.

Background: Plant cell culture technology is a potential way to produce polyphenols, however, this way is still trapped in the dilemma of low content and yield. Elicitation is regarded as one of the most effective ways to improve the output of the secondary metabolites, and therefore has attracted extensive attention. Methods: Five elicitors including 5-aminolevulinic acid (5-ALA), salicylic acid (SA), methyl jasmonate (MeJA), sodium nitroprusside (SNP) and Rhizopus Oryzae Elicitor (ROE) were used to improve the content and yield of polyphenols in the cultured Cyclocarya paliurus (C. paliurus) cells, and a co-induction technology of 5-ALA and SA was developed as a result. Meanwhile, the integrated analysis of transcriptome and metabolome was adopted to interpret the stimulation mechanism of co-induction with 5-ALA and SA. Results: Under the co-induction of 50 μM 5-ALA and SA, the content and yield of total polyphenols of the cultured cells reached 8.0 mg/g and 147.12 mg/L, respectively. The yields of cyanidin-3-O-galactoside, procyanidin B1 and catechin reached 28.83, 4.33 and 2.88 times that of the control group, respectively. It was found that expressions of TFs such as CpERF105, CpMYB10 and CpWRKY28 increased significantly, while CpMYB44 and CpTGA2 decreased. These great changes might further make the expression of CpF3'H (flavonoid 3'-monooxygenase), CpFLS (flavonol synthase), CpLAR (leucoanthocyanidin reductase), CpANS (anthocyanidin synthase) and Cp4CL (4-coumarate coenzyme A ligase) increase while CpANR (anthocyanidin reductase) and CpF3'5'H (flavonoid 3', 5'-hydroxylase) reduce, ultimately enhancing the polyphenols accumulation Conclusion: The co-induction of 5-ALA and SA can significantly promote polyphenol biosynthesis in the cultured C. paliurus cells by regulating the expression of key transcription factors and structural genes associated with polyphenol synthesis, and thus has a promising application.

Microbial and clinical epidemiology of invasive fungal rhinosinusitis in hospitalized COVID-19 patients, the divergent causative agents.

Since COVID-19 spread worldwide, invasive fungal rhinosinusitis (IFRS) has emerged in immunocompromised patients as a new clinical challenge. In this study, clinical specimens of 89 COVID-19 patients who presented clinical and radiological evidence suggestive of IFRS were examined by direct microscopy, histopathology, and culture, and the isolated colonies were identified through DNA sequence analysis. Fungal elements were microscopically observed in 84.27% of the patients. Males (53.9%) and patients over 40 (95.5%) were more commonly affected than others. Headache (94.4%) and retro-orbital pain (87.6%) were the most common symptoms, followed by ptosis/proptosis/eyelid swelling (52.8%), and 74 patients underwent surgery and debridement. The most common predisposing factors were steroid therapy (n=83, 93.3%), diabetes mellitus (n=63, 70.8%), and hypertension (n=42, 47.2%). The culture was positive for 60.67% of the confirmed cases, and Mucorales were the most prevalent (48.14%) causative fungal agents. Different species of Aspergillus (29.63%) and Fusarium (3.7%) and a mix of two filamentous fungi (16.67%) were other causative agents. For 21 patients, no growth was seen in culture despite a positive result on microscopic examinations. In PCR-sequencing of 53 isolates, divergent fungal taxons, including 8 genera and 17 species, were identified as followed: Rhizopus oryzae (n=22), Aspergillus flavus (n=10), A. fumigatus (n=4), A. niger (n=3), R. microsporus (n=2), Mucor circinelloides, Lichtheimia ramosa, Apophysomyces variabilis, A. tubingensis, A. alliaceus, A. nidulans, A. calidoustus, Fusarium fujikuroi/proliferatum, F. oxysporum, F. solani, Lomentospora prolificans, and Candida albicans (each n=1). In conclusion, a diverse set of species involved in COVID-19-associated IFRS was observed in this study. Our data encourage specialist physicians to consider the possibility of involving various species in IFRS in immunocompromised and COVID-19 patients. In light of utilizing molecular identification approaches, the current knowledge of microbial epidemiology of invasive fungal infections, especially IFRS, may change dramatically.

Metatranscriptomics Reveals Sequential Expression of Genes Involved in the Production of Melanogenesis Inhibitors by the Defined Microbial Species in Fermented Unpolished Black Rice.

Fermented products require metabolic enzymes from the microbial community for desired final products. Using a metatranscriptomic approach, the role of microorganisms in fermented products on producing compounds with a melanogenesis inhibition activity has not yet been reported. Previously, unpolished black rice (UBR) fermented with the E11 starter containing Saccharomyces cerevisiae, Saccharomycopsis fibuligera, Rhizopus oryzae, and Pediococcus pentosaceus (FUBR) showed potent melanogenesis inhibition activity. This study aimed to investigate the function of these defined microbial species in producing melanogenesis inhibitors in the FUBR using a metatranscriptomic approach. The melanogenesis inhibition activity increased in a fermentation time-dependent manner. Genes related to melanogenesis inhibitors synthesis such as carbohydrate metabolism, amino acids synthesis, fatty acids/unsaturated fatty acids synthesis, and carbohydrate transporters were analyzed. Most genes from R. oryzae and P. pentosaceus were upregulated in the early stage of the fermentation process, while those of S. cerevisiae and S. fibuligera were upregulated in the late stage. FUBR production using different combinations of the four microbial species shows that all species were required to produce the highest activity. The FUBR containing at least R. oryzae and/or P. pentosaceus exhibited a certain level of activity. These findings were in agreement with the metatranscriptomic results. Overall, the results suggested that all four species sequentially and/or coordinately synthesized metabolites during the fermentation that led to a FUBR with maximum melanogenesis inhibition activity. This study not only sheds light on crucial functions of certain microbial community on producing the melanogenesis inhibitors, but also paves the way to initiate quality improvement of melanogenesis inhibition activity in the FUBR. IMPORTANCE Fermentation of food is a metabolic process through the action of enzymes from certain microorganisms. Although roles of the microbial community in the fermented food were investigated using metatranscriptomic approach in terms of flavors, but no study has been reported so far on the function of the microorganisms on producing compounds with a melanogenesis inhibition activity. Therefore, this study explained the roles of the defined microorganisms from the selected starter in the fermented unpolished black rice (FUBR) that can produce melanogenesis inhibitor(s) using metatranscriptomic analysis. Genes from different species were upregulated at different fermentation time. All four microbial species in the FUBR sequentially and/or coordinately synthesized metabolites during fermentation that led to a FUBR with maximal melanogenesis inhibition activity. This finding contributes to a deeper understanding of the roles of certain microbial community during fermentation and led to the knowledge-based improvement for the fermented rice with potent melanogenesis inhibition activity.

Photodynamic inactivation of Rhizopus oryzae – in vitro study

Photodynamic inactivation of Rhizopus oryzae – in vitro study

Photodynamic inactivation of Rhizopus oryzae – in vitro study

Photodynamic inactivation of Rhizopus oryzae – in vitro study

Glutaraldehyde-crosslinked Rhizopus oryzae whole cells show improved catalytic performance in alkene epoxidation

BackgroundExisting methods for alkene epoxidation are based on lipase-catalysed perhydrolysis. However, the inactivation of the expensive lipase enzyme is problematic for enzymatic epoxidation at large scales due to the use of hydrogen peroxide and peracids at high concentrations in the reaction. The immobilisation of whole cells appears to be a promising approach to alleviate this problem.ResultsA green oxidation system containing hydrogen peroxide, Na3C6H5O7, an acyl donor, and glutaraldehyde (GA)-crosslinked cells of Rhizopus oryzae was developed for the epoxidation of alkenes. GA-crosslinked cells of Rhizopus oryzae were adopted as a biocatalyst into the epoxidation system. A variety of alkenes were oxidised with this system, with a 56–95% analytical yield of the corresponding epoxides. The catalytic performance of the crosslinked treated cells was substantially improved compared to that of the untreated cells and the initial reaction rate increased from 126.71 to 234.72 mmol/L/h, retaining 83% yields even after four batches of reactions. The addition of 3.5 mmol Na3C6H5O7 not only acts as an acid-trapping reagent to eliminate the negative effect of the carboxylic acid on the alkene oxide but also forms a saturated salt solution with the aqueous phase, affecting the concentration of H2O2 in the three phases and thus the epoxidation reaction. Organic solvents with a logP value > 0.68 were good at producing hydroxy peracids; however, this method is only suitable for oxidation in a two-liquid phase.ConclusionsCompared with other lipase biocatalysts, the GA-crosslinked whole-cell biocatalyst is inexpensive, readily available, and highly stable. Therefore, it can be considered promising for industrial applications.

Changes in polyphenols composition and antioxidative properties of hemp (Cannabis sativa L.) inflorescences pretreated by Ganoderma lucidum

The purpose of present study was to improve the antioxidative activity and explore the potential biotransformation mechanism in hemp (Cannabis sativa L.) inflorescences pretreated with Ganoderma lucidum. Ganoderma lucidum, Rhizopus oryzae and Aspergillus oryzae were applied to the transformation of bioactive substances in hemp inflorescences, and the results showed that G. lucidum was the most suitable strain for pretreatment of hemp inflorescences.The optimum time and temperature of G. lucidum pretreatment was 28 ℃ and 7 days respectively. Under the optimum conditions, the 1,1-diphenyl-2-picryl-hydrazyl (DPPH·) and hydroxyl radical (-OH) scavenging ratio were increased by 1.73 and 1.46-fold respectively than that of untreated hemp inflorescences. The total polyphenols (TP) and total flavonoids (TF) were significantly decreased. The metabonomic analysis indicated that, after pretreatment, flavonoids were methoxylated to O-methylated flavonoids. In addition, cinnamic acids hydroxylated to hydroxycinnamic acids. Moreover, flavonoid O-glycosides were rapidly metabolized to release aglycones. Our results showed that pretreatment by G. lucidum exerts significant changes on the polyphenols composition of hemp inflorescences, thus modifying their antioxidative properties.

Rhizopus oryzae Inulinase Production and Characterization with Application in Chicory Root Saccharification

The objective of this study was to create a fermentation process for the production of inulinase, an important enzyme with numerous applications in the food and pharmaceutical industries, using low-cost agricultural waste as substrates for Rhizopus oryzae NRRL 3563. High titer inulinase production in chicory roots by Rhizopus oryzae in a submerged culture was accomplished using a statistical experimental design. A two-level Plackett–Burman design followed by a three-level Box–Behnken design producing a high inulinase titer of 1085.11 U/mL, 2.83-fold the maximum level, was obtained in the screening experiment. The optimal levels were as follows: chicory root, 10 g/L; NaNO3, 5 g/L; and KCl, 0.2 g/L. The produced inulinase enzyme was purified using 70% ammonium sulfate precipitation and ultra-filtration causing 3.63-fold purification with 60% activity recovery. The enzyme had a molecular weight of approximately 130 KDa. The purified enzyme showed optimum activity at 50 °C and pH 6.0. The pH stability range was three to six and the temperature stability was up 70 °C. The purified inulinase could hydrolyze inulin and sucrose, but not cellobiose or soluble starch. Km and Vmax for inulin were determined to be 0.8 mg/mL and 50,000 U/mg, respectively. The two-level Plackett–Burman design was applied followed by a Box–Behnken model for optimization of fermentation conditions. Accordingly, the optimal combination of fermentation was a reaction time of seven hours, a temperature of 60 °C, and an enzyme concentration of 40,000 U/mL, which resulted in a 58.07% saccharification yield. The characteristics of the enzyme and its kinetic parameters suggested that it was highly effective in the fermentation of inulin and inulin-containing substrates. Additionally, it raises the potential of using inulinase enzymes in pharmaceutical and food industries.

Exploring the potential of Rhizopus oryzae AUMC14899 as a novel endophytic fungus for the production of l-tyrosine and its biomedical applications

BackgroundA significant threat to the public's health is the rise in antimicrobial resistance among numerous nosocomial bacterial infections. This may be a detriment to present initiatives to enhance the health of immune-compromised patients. Consequently, attention has been devoted to exploring new bioactive compounds in the field of drug discovery from endophytes. Therefore, this study is the first on the production of l-tyrosine (LT) as a promising bio-therapeutic agent from endophytic fungi.ResultsA new endophytic fungal isolate has been identified for the first time as Rhizopus oryzae AUMC14899 from Opuntia ficus-indica (L.) and submitted to GenBank under the accession number MZ025968. Separation of amino acids in the crude extract of this fungal isolate was carried out, giving a higher content of LT, which is then characterized and purified. LT exhibited strong antibacterial and anti-biofilm activities against multidrug-resistant Gram-negative and Gram-positive bacteria. The recorded minimum inhibitory concentration (MIC) values ranged from 6 to 20 µg/ml. In addition, LT caused a strong reduction in biofilm formation and disrupted the preformed biofilm. Moreover, results indicated that LT supported cell viability, evidencing hemocompatibility and no cytotoxicity.ConclusionOur findings suggest that LT has potential as a therapeutic agent due to its potential antibacterial, anti-biofilm, hemocompatibility, and lack of cytotoxic activities, which may also increase the range of therapy options for skin burn infections, leading to the development of a novel fungal-based drug.

Disease Duration and Chronic Complications Associate With Immune Activation in Individuals With Longstanding Type 1 Diabetes.

Type 1 diabetes (T1D) is associated with alterations of the immune response which persist even after the autoimmunity aspect is resolved. Clinical factors that cause dysregulation, however, are not fully understood. To identify clinical factors that affect immune dysregulation in people with longstanding T1D. In this cross-sectional study, 243 participants with longstanding T1D were recruited between February 2016 and June 2017 at the Radboudumc, the Netherlands. Blood was drawn to determine immune cell phenotype and functionality, as well as circulating inflammatory proteome. Multivariate linear regression was used to determine the association between glycated hemoglobin (HbA1c) levels, duration of diabetes, insulin need, and diabetes complications with inflammation. HbA1c level is positively associated with circulating inflammatory markers (P < .05), but not with immune cell number and phenotype. Diabetes duration is associated with increased number of circulating immune cells (P < .05), inflammatory proteome (P < .05), and negatively associated with adaptive immune response against Mycobacterium tuberculosis and Rhizopus oryzae (P < .05). Diabetes nephropathy is associated with increased circulating immune cells (P < .05) and inflammatory markers (P < .05). Disease duration and chronic complications associate with persistent alterations in the immune response of individuals with long standing T1D.

The Impact of Microorganisms on the Performance of Linseed Oil and Tung Tree Oil Impregnated Composites Made of Hemp Shives and Corn Starch

In this study, the performance characteristics of hemp shives impregnated with linseed oil and tung tree oil (HS)- and corn starch (CS)-based biocomposites containing flame retardants were evaluated before and after treatment with the mixture of bacterium Pseudomonas putida and fungus Rhizopus oryzae. Enzymatic activities and physical-mechanical properties such as water absorption, thickness swelling, compressive strength, and thermal conductivity were tested to evaluate the suitability of selected composites for thermal insulation purposes. In addition, electron microscopy was used to investigate the impact of microorganisms on the microstructure of the material. It was determined that the type of oil used for impregnation significantly affects the properties of biocomposites after 6 months of incubation with mixture of bacterium P. putida and fungus Rh. oryzae. Biocomposites impregnated with linseed oil and after treatment with a mixture of microorganisms had cellulase activity of 25 U/mL, endo β-1-4-glucanase activity of 26 U/mL, lipase activity of 101 U/mL, only a 10% decrease in compressive strength, 50% higher short-term water absorption, unchanged swelling in thickness, and slightly decreased thermal conductivity compared to control biocomposites. At the same time, biocomposites with tung tree oil had a much more pronounced deterioration of the properties tested, cellulase activity of 28 U/mL, endo β-1-4-glucanase activity of 37 U/mL, lipase activity of 91 U/mL, two times lower compressive strength and two times higher short-term water absorption, 2.5 times greater thickness swelling, and a slightly increased thermal conductivity. We conclude that linseed oil provides better protection against the action of microorganisms compared to impregnation with tung tree oil.

Glucose assimilation rate determines the partition of flux at pyruvate between lactic acid and ethanol in Saccharomyces cerevisiae.

Engineered Saccharomyces cerevisiae expressing a lactic acid dehydrogenase can metabolize pyruvate into lactic acid. However, three pyruvate decarboxylase (PDC) isozymes drive most carbon flux toward ethanol rather than lactic acid. Deletion of endogenous PDCs will eliminate ethanol production, but the resulting strain suffers from C2 auxotrophy and struggles to complete a fermentation. Engineered yeast assimilating xylose or cellobiose produce lactic acid rather than ethanol as a major product without the deletion of any PDC genes. We report here that sugar flux, but not sensing, contributes to the partition of flux at the pyruvate branch point in S. cerevisiae expressing the Rhizopus oryzae lactic acid dehydrogenase (LdhA). While the membrane glucose sensors Snf3 and Rgt2 did not play any direct role in the option of predominant product, the sugar assimilation rate was strongly correlated to the partition of flux at pyruvate: fast sugar assimilation favors ethanol production while slow sugar assimilation favors lactic acid. Applying this knowledge, we created an engineered yeast capable of simultaneously converting glucose and xylose into lactic acid, increasing lactic acid production to approximately 17gL-1 from the 12gL-1 observed during sequential consumption of sugars. This work elucidates the carbon source-dependent effects on product selection in engineered yeast.

In Vitro Evaluation of the Antagonism of Saprophyte and Endophytic Fungi Isolated from Groundnut (Arachis hypogaea) Against Soil-Transmitted Diseases Sclerotium rolfsii

Certain soil-borne diseases can have a detrimental effect on the quality and quantity of peanut production. One of the soil-borne diseases is Sclerotium rolfsii, a fungus that is the primary limiting factor in peanut plants. An alternative to controlling this fungus is to use saprophytic and endophytic fungi. The purpose of this study was to evaluate several saprophytic and endophytic fungi isolated from peanut plants in vitro as antagonistic agents against S. rolfsii. Isolation of saprophytic and endophytic fungi was carried out using the stratified dilution method. The fungal antagonism test was carried out using a non-factorial completely randomized design with 3 replications. The parameters observed in this study included the inhibiting zone, the diameter of the isolate colonies, the growth area, and the interaction of saprophytic and endophytic fungi with S. rolfsii. Five families and eight species of soil fungi were identified using macroscopic and microscopic identification techniques. The type of soil fungus has a large impact on the growth rate and inhibition area. Mucor hiemalis grew at the fastest rate, while Rhizopus oryzae had the largest inhibition zone.

First Report of Rhizopus arrhizus (syn. R. oryzae) Causing Garlic Bulb Soft Rot in Hebei Province, China.

Rhizopus soft rot occurs on the succulent tissues of vegetables, fruits, and ornamental plants throughout the world (Cui et al. 2019). When the garlic is in the seedling stage in the fields (Fig. S1) in November 2021, a disease outbreak on garlic bulbs suspected as Rhizopus soft rot occurred in Daming County, Handan City, Hebei Province of China (N 36°17', E 115° 13'). This disease symptom was first found in the garlic seedling stage in China. Disease incidence was 10% to 30% in cultivated garlic bulbs. There were soft water-soaked lesions on the surface of diseased garlic bulbs and the interiors were brown and soft. In the disease severe field, white to gray mycelia were observed on the diseased garlic bulbs. Infected garlic bulbs were sampled to isolate and determine the identity of the disease-causing organism. Symptomatic bulbs were surface sterilized with 1% NaClO for 2 min, dipped in 75% ethanol for 3 min and rinsed three times with autoclaved distilled water. Small pieces of the inner decayed tissue were removed and cultured on potato dextrose agar (PDA) at 28°C for 2 to 3 days. Five white colonies grew on PDA and then they became brownish gray to blackish-gray mycelium. The fungal strains were purified by hyphal-tip isolation method. To determine the identity of the five isolated fungi, we analyzed their internal transcribed spacer (ITS) region sequences (Jung et al. 2012). BLAST analysis of the ITS sequences from DSF-0-2 (accession no. ON706022), DSF-0-3 (accession no. ON706021), DSF-0-4 (accession no. ON706020), DSF-0-5 (accession no. ON706019) and DSF-0-6 (accession no. ON706018) were all 100% identical with Rhizopus arrhizus (syn. Rhizopus oryzae). Phylogenetic trees were constructed using the neighbor-joining method of MEGA11 based on the sequences of ITS rRNA gene (Walther et al. 2013). Phylogenetic trees indicated that isolates were most likely Rhizopus arrhizus (syn. Rhizopus oryzae) (Fig. S2). We selected one isolated strain, DSF-0-2, for characterize the morphology and test its ability to cause garlic bulb soft rot. Under the microscope, nonseptate rhizoids, sporangia, and sporangiospores were observed (Fig. S1). Sporangiospores were unequal, subglobose, numerous irregular, or oval, and 9.7 (6.2 - 12.5) × 6.5 (4.1 - 8.5) μm (n = 50) in diameter. The sporangia were globose, black, 121.5 (65 - 198) μm (n = 50) in diameter. Based on the rDNA-ITS sequencing and the morphological characteristics, the DSF-0-2 isolate was identified as Rhizopus arrhizus (syn. Rhizopus oryzae) (Zheng et al. 2007; Abeywickrama et al. 2020). To complete Koch's postulates, surface-sterilized healthy garlic bulbs were inoculated with R. arrhizus isolate DSF-0-2. A 1.0-ml sterile syringe was used to inject 50 μl of a 106 conidia/ml suspension into each of five healthy bulbs. As a control, garlic bulbs were treated with sterile distilled water. The inoculated and control bulbs were incubated at 28°C for 7 days. The bulbs inoculated with R. arrhizus DSF-0-2 showed symptoms of water soaking, and the tissues were brown and soft throughout the bulb at 7 days (Fig. S1). Results of the three trials were the same. No symptoms were observed in the control group. R. arrhizus was reisolated from the symptomatic garlic bulb and confirmed as such based-on colony and sporangia morphology and ITS sequence. There were some reports that R. arrhizus infects cassava tubers and potato tubers (Amadioha and Markson 2007; Cui et al. 2019). To our knowledge, this is the first report of R. arrhizus (syn. Rhizopus oryzae) associated with soft rot on garlic bulb in the seedling stage in China. This disease has posed a potential threat during garlic seedling stage in the field. Management measures should be considered before this disease outbreaks widely. Garlic bulbs died in the seedling stage, which caused production reduction, serious economic loss and soil pollution. This finding may help to take effective control measures for this disease.

Molecular detection and identification of fungal pathogens infections occurring in COVID-19 recovered patients.

The major outbreak of Corona virus disease COVID-19 caused by SARS-CoV-2 had brought about 4.55million deaths and had shaken the health care system all over the world. From the year 2020 the recovered COVID-19patients had started to develop microbial infection, most predominantly fungal infection in which Mucormycosis gained immediate attention as it has worsen the mortality rate in humans. In the present study of 53COVID-19recovered patients presented with microbial infection, the analysis of frequency distribution of fungal infection preponderantly with Rhizopus oryzae, followed by Aspergillus and Candida species.

Effects of Lipase Immobilization Conditions and Support Materials for the Production of Structured Triacylglycerols

AbstractStructured triacylglycerols (STAG) with desired properties can be synthesized by transesterification using immobilized lipases. Herein, the effect of immobilization conditions and support material on the immobilization yield, specific activity and regioselectivity of lipases from Rhizomucor miehei (RML) and Rhizopus oryzae (ROL) in the production of STAG, are evaluated. Four different support materials utilizing adsorption and one with covalent binding are investigated. Ammonium sulfate is found to significantly increase the activity‐based immobilization yield (12% and 38% for RML and ROL, respectively) and specific activity on Accurel MP1000 (MP1000) when used as the immobilization buffer. Furthermore, the immobilization principle and support material influenced both the activity and regiospecificity. Immobilization by adsorption is found to result in higher catalytic activity, while covalent binding resulted in lipase inactivation. For RML, the highest specific activity of 43 µmol STAG min−1 g−1 (U) is obtained on MP1000, while ROL, which exhibited higher activities in general, results in a maximum activity of 120 U on Lifetech ECR8806. The obtained specific activites are comparable to the commercial preparations Novozym 40086 (45 U) and Lipase DF “Amano” IM (147 U) while the regiospecificity of the developed preparations is even higher, forming at least 64% less byproduct.Practical applications: There is an increasing need for lipids with specific nutritional and physical properties in health, nutrition, and food applications. In this context, STAG are highly promising products due to the possibility to tailor the composition to obtain the desired properties. Immobilized lipases are the catalyst of choice for the production of STAG, in which activity and regioselectivity are particularly important parameters for the process performance. In this study, it is shown that these parameters can be affected by the immobilization conditions and support material. Immobilized preparations with high activity and excellent regiospecificity are created on commercially available supports. This shows the possibility of STAG synthesis with high purity and beneficial properties.

Effects of Combined Pure Cultures of Rhizopus sp. (Rhizopus oryzae, Rhizopus oligosporus, and Rhizopus stolonifer) on Tempeh Extract Yogurt as a Functional Food

Background: Tempeh Extract Yogurt was an alternative to tempeh product development that can be used as a functional product. This study aimed to determine the functional food potential of yogurt with tempeh extract by including tempeh using mixed cultures. Methods: The ratio of making tempeh Rhizopus oryzae, Rhizopus oligosporus, Rhizopus stolonifer inoculum of 1:1:1, 1:2:1, 2:1:2, 1:1:2, 2:1:1, respectively; RAPRIMA tempeh starter as a control group was fermented with Lactic Acid Bacteria to produce tempeh extract yogurt. Results: Tempeh extract yogurt by SNI 2891:2009 with the characteristic’s thick liquid, specific aroma, sour taste, and homogeneous consistency. The highest form of vitamin B12 and folate in Tempeh Extract Yogurt was on a 1:2:1 ratio (0.072 g/100 mL and 0.059 g/100 mL), and the genistein isoflavone in RAPRIMA was 3.669 g/100 mL with a pH value of 4.2 and a total bacterium of 14,3 x 108 CFU/mL. Conclusion: Tempeh extract yogurt contains several vitamin B12, folate, isoflavones, and total bacteria Tempeh Extract Yogurt be responsible as a functional food.

New neuroprotective sesquiterpene lactate esters from carotol biotransformation

Biotransformation of carotol (1), the major sesquiterpenoid of carrot fruit essential oil, by Rhizopus oryzae yielded two new lactate derivatives: 9α-hydroxydaucol-9-lactate (2) and 9α-hydroxydaucol-3,9-dilactate (3). The structures of 1-3 were characterized by 1 & 2D NMR spectroscopy in conjunction with HR-ESIMS. Cholinergic deficiency and neuroinflammation are key detrimental processes involved in the etiology of Alzheimer's disease (AD). This study demonstrated that compounds 1-3 in addition to the essential oil showed considerable in-vitro inhibition of cyclooxygenase-2 (COX-2), 5-lipoxygenase (5-LOX), and butyrylcholinesterase (BChE). Metabolite 2 showed improved inhibitory activities (9.03, 0.56, and 0.64 μM) against COX-2, 5-LOX, and BChE enzymes, compared to the parent compound, carotol 1 (82.57, 2.11, and 0.74 μM, respectively). The results were correlated to the in-silico binding interactions with the proteins of these enzymes by molecular docking. Carrot essential oil and carotol derivatives are suggested by this study as a potential treatment for neurodegenerative disease associated with neuroinflammation.

Antifungal and Nanozyme Activities of Metal–Organic Framework‐derived CuO@C

Postharvest fungal infection is the common cause of fresh fruit losses. Thus, effective antifungal agents are highly required. Herein, copper‐based metal–organic frameworks (MOFs) were used as a precursor for the synthesis of copper oxide (CuO) embedded into carbon (CuO@C). The materials' crystallinity was confirmed using X‐ray diffraction. X‐ray photoelectron spectroscopy (XPS) and atomic absorption spectroscopy (AAS) approve the elemental species and concentration of copper in the synthesized materials. Transmission electron microscopy (TEM), high‐resolution TEM (HR‐TEM), and scanning electron microscopy (SEM) reveal the successful formation of CuO‐embedded C, that is, CuO@C, with a particle size of 36–123 nm. CuO@C nanoparticles are used as an antifungal agent against four economic postharvest plant pathogens; Alternaria alternata (black spot disease of tomato), Fusarium oxysporum (rot disease of tomato), Penicillium digitatum (gray mold disease of lemon), and Rhizopus oryzae (soft rot disease of banana). The inhibition zones were 36 ± 0.5, 20.2 ± 0.4, 16 ± 0.5, and 10.2 ± 0.1 mm for R. oryzae, P. digitatum, F. oxysporum, and A. alternata, respectively. CuO@C antifungal action includes significant inhibition of cellulases and amylase enzyme activity, decreasing the potentiality of fungal pathogenicity and fruit disease. CuO@C exhibited high antifungal activity against postharvest antifungal species.