A smartphone-combined ratiometric fluorescence molecularly imprinted probe based on biomass-derived carbon dots for determination of tyramine in fermented meat products.

A ratiometric fluorescence molecularly imprinted probe employing two distinct emission wavelengths of biomass carbon dots was developed for highly selective and visual quantitative detection of tyramine in fermented meat products. The red emission biomass carbon dots were employed as responsive elements, and the blue ones were utilized as the reference elements. The molecularly imprinted polymers were incorporated in the ratiometric sensing to distinguish and adsorb tyramine. With the linear range of 1-60 µg/L, the ratiometric fluorescence molecularly imprinted probe was successfully applied to detect tyramine in real samples with the satisfactory recoveries of 79.74-112.12% and the detect limitation of 1.3 µg/kg, indicating that this probe has great potential applications for the detection of tyramine in real samples. Moreover, smartphone-based fluorescence signal recognition analysis on hand has been developed for the quantitative analysis of tyramine, providing a portable visual optical analysis terminal for rapid on-site determination of tyramine.

Catalytic Synthesis of (S)-CHBE by Directional Coupling and Immobilization of Carbonyl Reductase and Glucose Dehydrogenase.

Ethyl (S)-4-chloro-3-hydroxybutyrate ((S)-CHBE) is an important chiral intermediate in the synthesis of the cholesterol-lowering drug atorvastatin. Studying the use of SpyTag/SpyCatcher and SnoopTag/SnoopCatcher systems for the asymmetric reduction reaction and directed coupling coenzyme regeneration is practical for efficiently synthesizing (S)-CHBE. In this study, Spy and Snoop systems were used to construct a double-enzyme directed fixation system of carbonyl reductase (BsCR) and glucose dehydrogenase (BsGDH) for converting 4-chloroacetoacetate (COBE) to (S)-CHBE and achieving coenzyme regeneration. We discussed the enzymatic properties of the immobilized enzyme and the optimal catalytic conditions and reusability of the double-enzyme immobilization system. Compared to the free enzyme, the immobilized enzyme showed an improved optimal pH and temperature, maintaining higher relative activity across a wider range. The double-enzyme immobilization system was applied to catalyze the asymmetric reduction reaction of COBE, and the yield of (S)-CHBE reached 60.1% at 30 °C and pH 8.0. In addition, the double-enzyme immobilization system possessed better operational stability than the free enzyme, and maintained about 50% of the initial yield after six cycles. In summary, we show a simple and effective strategy for self-assembling SpyCatcher/SnoopCatcher and SpyTag/SnoopTag fusion proteins, which inspires building more cascade systems at the interface. It provides a new method for facilitating the rapid construction of in vitro immobilized multi-enzyme complexes from crude cell lysate.

Characterizations of biogenic selenium nanoparticles and their anti-biofilm potential against Streptococcus mutans ATCC 25175.

INTRODUCTION: S. mutans has been identified as the primary pathogenic bacterium in biofilm-mediated dental caries. The biogenic selenium nanoparticles (SeNPs) produced by L. plantarum KNF-5 were used in this study against S. mutans ATCC 25175. OBJECTIVES: The aims of this study were: (1) the biosynthesis of SeNPs by L. plantarum KNF-5, (2) the characterization of SeNPs, (3) the investigation of the inhibitory effect of biogenic SeNPs against S. mutans ATCC 25175, and (4) the determination of the anti-biofilm potential of SeNPS against S. mutans ATCC 25175. METHODOLOGY: 3 mL of the culture was added to 100 mL of MRS medium and incubated. After 4 h, Na2SeO3 solution (concentration 100 µg/mL) was added and incubated at 37 °C for 36 h. The color of the culture solution changed from brownish-yellow to reddish, indicating the formation of SeNPs. The characterization of SeNPs was confirmed by UV-Vis spectrophotometry, FTIR, SEM-EDS and a particle size analyzer. The antibacterial activity was determined by the disk diffusion method, the MIC by the micro-double dilution method, and the biofilm inhibitory potential by the crystal violet method and the MTT assay. The effect of SeNPs on S. mutans ATCC 25175 was determined using SEM and CLSM spectrometry techniques. The sulfate-anthrone method was used to analyze the effect of SeNPs on insoluble extracellular polysaccharides. The expression of genes in S. mutans ATCC 25175 was analyzed by real-time quantitative polymerase chain reaction (RT-qPCR). PREPARATION OF NANOPARTICLES: SeNPs produced by probiotic bacteria are considered a safe method. In this study, L. plantarum KNF-5 (probiotic strain) was used for the production of SeNPs. RESULTS: The biogenic SeNPs were spherical and coated with proteins and polysaccharides and had a diameter of about 270 nm. The MIC of the SeNPs against S. mutans ATCC 25175 was 3.125 mg/mL. Biofilm growth was also significantly suppressed at this concentration. The expression of genes responsible for biofilm formation (GtfB, GtfC, BrpA and GbpB,) was reduced when S. mutans ATCC 25175 was treated with SeNPs. CONCLUSION: It was concluded that the biogenic SeNPs produced by L. plantarum KNF-5 was highly effective to inhibit the growth of S. mutans ATCC 25175. NOVELTY STATEMENT: The application of biogenic SeNPs, a natural anti-biofilm agent against S. mutans ATCC 25175. In the future, this study will provide a new option for the prevention and treatment of dental caries.

Study of selenium enrichment metabolomics in Bacillus subtilis BSN313 via transcriptome analysis.

In this study, the transcriptome analysis was practiced to identify potential genes of probiotic Bacillus subtilis BSN313 involved in selenium (Se) enrichment metabolism. The transcriptomic variation of the strain was deliberated in presence of three different sodium selenite concentrations (0, 3, and 20 µg/mL). The samples were taken at 1 and 13 h subsequent to inoculation of selenite and gene expression profiles in Se metabolism were analyzed through RNA sequencing. The gene expression levels of the pre log phase were lower than the stationary phase. It is because, the bacteria has maximum grown with high concentration of Se (enriched with organic Se), at stationary phase. Bacterial culture containing 3 µg/mL concentration of inorganic Se (sodium selenite) has shown highest gene expression as compared to no or high concentration of Se. This concentration (3 µg/mL) of sodium selenite (as Se) in the medium promoted the upregulation of thioredoxin reductase expression, whereas its higher Se concentration inhibited the formation of selenomethionine (SeMet). The result of 5 L bioreactor fermentation showed that SeMet was also detected in the fermentation supernatant as the growth entered in the late stationary phase and reached up to 857.3 ng/mL. The overall intracellular SeMet enriched content in BSN313 was extended up to 23.4 µg/g dry cell weight. The other two selenoamino acids (Se-AAs), methyl-selenocysteine, and selenocysteine were hardly detected in medium supernatant. From this study, it was concluded that SeMet was the highest content of organic Se byproduct biosynthesized by B. subtilis BSN313 strain in Se-enriched medium during stationary phase. Thus, B. subtilis BSN313 can be considered a commercial probiotic strain that can be used in the food and pharmaceutical industries. This is because it can meet the commercial demand for Se-AAs (SeMet) in both industries.

Expression of BSN314 lysozyme genes in Escherichia coli BL21: a study to demonstrate microbicidal and disintegarting potential of the cloned lysozyme.

This study is an extension of our previous studies in which the lysozyme was isolated and purified from Bacillus subtilis BSN314 (Naveed et al., 2022; Naveed et al., 2023). In this study, the lysozyme genes were cloned into the E. coli BL21. For the expression of lysozyme in E. coli BL21, two target genes, Lyz-1 and Lyz-2, were ligated into the modified vector pET28a to generate pET28a-Lyz1 and pET28a-Lyz2, respectively. To increase the production rate of the enzyme, 0.5-mM concentration of IPTG was added to the culture media and incubated at 37 °C and 220 rpm for 24 h. Lyz1 was identified as N-acetylmuramoyl-L-alanine amidase and Lyz2 as D-alanyl-D-alanine carboxypeptidase. They were purified by multi-step methodology (ammonium sulfate, precipitation, dialysis, and ultrafiltration), and antimicrobial activity was determined. For Lyz1, the lowest MIC/MBC (0.25 µg/mL; with highest ZOI = 22 mm) were recorded against Micrococcus luteus, whereas the highest MIC/MBC with lowest ZOI were measured against Salmonella typhimurium (2.50 µg /mL; with ZOI = 10 mm). As compared with Aspergillus oryzae (MIC/MFC; 3.00 µg/mL), a higher concentration of lysozyme was required to control the growth of Saccharomyces cerevisiae (MIC/MFC; 50 µg/mL). Atomic force microscopy (AFM) was used to analyze the disintegrating effect of Lyz1 on the cells of selected Gram-positive bacteria, Gram-negative bacteria, and yeast. The AFM results showed that, as compared to Gram-negative bacteria, a lower concentration of lysozyme (Lyz1) was required to disintegrate the cell of Gram-positive bacteria.

Nutritional composition assessment and antimicrobial activity of Catostylus perezi, jellyfish blooms along the coast of Pakistan: an awareness to avoid food neophobia in Pakistan.

This study highlighted the nutritional importance of Catostylus perezi (an edible jellyfish) in Pakistan; a society where a large proportion of the population suffers from malnutrition, while C. perezi, a blessing of the sea, is wasted or exported. In the present study, the amino acid and fatty acid profiles of the oral arms and umbrella of C. perezi were determined by HPLC. The total amino acid concentration (ΣAA) in the oral arms was 151.19 mg/100g, while in the umbrella it was 100.17 mg/100g. The ratio of total essential amino acids to total non-essential amino acids (TEAA/TNEAA) was 0.72 in the oral arms, while it was 0.70 in the umbrella. Higher amount of ω-3 with lower ratio of ω-6/ω-3 in oral arms (0.52), rather umbrella (ω-6/ω-3 ratio; 0.62). The antimicrobial activity, MIC, MBC, and MFC of the whole body of the edible jellyfish were determined. On the basis of polarity, different solvents were used, e.g. water, methanol, dichloromethane, chloroform, and n-hexane. Among all the extracts, the water extract gave the highest ZOI against C. xerosis (29 mm), while the n-hexane extract gave the lowest ZOI against S. aureus (MRSA) ATCC 33591 (8.20 mm). The water extract of C. perezi had high potential against C. xerosis with the highest AMI and PAI (1.53 and 153, respectively), while the same extract had the highest TAI against E. coli (81.43 mL/g). For fungi/yeast, the methanolic extract had the highest ZOI (29.70 mm) against S. cerevisiae and the lowest MIC/MFC (2.40 µg/mL) against the same pathogen. The n-Hexane extract gave the lowest ZOI (11.10 mm) against P. variotii and the highest MIC/MFC (31.60 µg/mL) against Penicillium sp. Atomic force microscopy (AFM) was used to analyse the disintegrating effect of the extracts (with the highest antimicrobial effect) on the cells of selected Gram-positive, Gram-negative and yeast species. The amino acid and fatty acid profiles and antimicrobial assessment showed that C. perezi has great nutritional importance, so the use of C. perezi as food is highly recommended for the Pakistani community.

Optimized methyl donor and reduced precursor degradation pathway for seleno-methylselenocysteine production in Bacillus subtilis.

BACKGROUND: Seleno-methylselenocysteine (SeMCys) is an effective component of selenium supplementation with anti-carcinogenic potential that can ameliorate neuropathology and cognitive deficits. In a previous study, a SeMCys producing strain of Bacillus subtilis GBACB was generated by releasing feedback inhibition by overexpression of cysteine-insensitive serine O-acetyltransferase, enhancing the synthesis of S-adenosylmethionine as methyl donor by overexpression of S-adenosylmethionine synthetase, and expressing heterologous selenocysteine methyltransferase. In this study, we aimed to improve GBACB SeMCys production by synthesizing methylmethionine as a donor to methylate selenocysteine and by inhibiting the precursor degradation pathway. RESULTS: First, the performance of three methionine S-methyltransferases that provide methylmethionine as a methyl donor for SeMCys production was determined. Integration of the NmMmt gene into GBACB improved SeMCys production from 20.7 to 687.4 µg/L. Next, the major routes for the degradation of selenocysteine, which is the precursor of SeMCys, were revealed by comparing selenocysteine hyper-accumulating and non-producing strains at the transcriptional level. The iscSB knockout strain doubled SeMCys production. Moreover, deleting sdaA, which is responsible for the degradation of serine as a precursor of selenocysteine, enhanced SeMCys production to 4120.3 µg/L. Finally, the culture conditions in the flasks were optimized. The strain was tolerant to higher selenite content in the liquid medium and the titer of SeMCys reached 7.5 mg/L. CONCLUSIONS: The significance of methylmethionine as a methyl donor for SeMCys production in B. subtilis is reported, and enhanced precursor supply facilitates SeMCys synthesis. The results represent the highest SeMCys production to date and provide insight into Se metabolism.

Selenium Enrichment of the Edible Medicinal Mushroom Antrodia camphorata by Submerged Fermentation.

Selenium (Se) is an essential nutrient element in human physiological metabolism and immune function. Supplementation of bioavailable Se will confer benefit on human life, especially when intake of this nutrient is inadequate. The edible and medicinal mushroom Antrodia camphorata is a unique fungus endemic to Taiwan, which has shown high therapeutic and nutritive value. This study is the first to demonstrate that A. camphorata can assimilate and transform sodium selenite into organic selenium. With an initial concentration of Se (IV) at 10 mg/L in 100 mL of the medium at 25 °C, the total selenium content in Se-enriched A. camphorata mycelia was 1281.3 ± 79.2 µg/g, in which the organic selenium content accounted for 88.1%. Further analysis demonstrated that selenium-enriched polysaccharide was the main form of Se present in A. camphorata (61.5% of the organic selenium). Four water-soluble Se-polysaccharide fractions were separated from A. camphorata, and ACP II was the major fraction of Se-polysaccharide. The scavenging efficiency of Se-polysaccharides on DPPH and ABTS radicals was determined, proving that selenium enrichment dramatically improved the in vitro antioxidant capacity of A. camphorata polysaccharide. Therefore, the selenium accumulation and transformation ability of A. camphorata provides an opportunity for developing this beneficent fungus into a novel selenium-enriched dietary or medicinal supplement.

Enhanced selenocysteine biosynthesis for seleno-methylselenocysteine production in Bacillus subtilis.

Seleno-methylselenocysteine (SeMCys) is an effective component for selenium supplementation with anti-carcinogenic potential and can ameliorate neuropathology and cognitive deficits. In this study, we aimed to engineer Bacillus subtilis 168 for the microbial production of SeMCys. First, the accumulation of intracellular selenocysteine (SeCys) as the precursor of SeMCys was enhanced through overexpression of serine O-acetyltransferase, which was desensitized against feedback inhibition by cysteine. Next, the S-adenosylmethionine (SAM) synthetic pathway was optimized to improve methyl donor availability through expression of S-adenosylmethionine synthetase. Further, SeMCys was successfully produced through expression of the selenocysteine methyltransferase in SeCys and SAM-producing strain. The increased expression level of selenocysteine methyltransferase benefited the SeMCys production. Finally, all the heterologous genes were integrated into the genome of B. subtilis, and the strain produced SeMCys at a titer of 18.4 µg/L in fed-batch culture. This is the first report on the metabolic engineering of B. subtilis for microbial production of SeMCys and provides a good starting point for future pathway engineering to achieve the industrial-grade production of SeMCys. KEY POINTS: • Expression of the feedback-insensitive serine O-acetyltransferase provided B. subtilis the ability of accumulating SeCys. • SAM production was enhanced through expressing S-adenosylmethionine synthetase in B. subtilis. • Expression of selenocysteine methyltransferase in SeCys and SAM-accumulating strain facilitated SeMCys production.

Purification, Characterization and Bactericidal Action of Lysozyme, Isolated from Bacillus subtillis BSN314: A Disintegrating Effect of Lysozyme on Gram-Positive and Gram-Negative Bacteria.

In the present study, lysozyme was purified by the following multi-step methodology: salt (ammonium sulfate) precipitation, dialysis, and ultrafiltration. The lysozyme potential was measured by enzymatic activity after each purification step. However, after ultrafiltration, the resulting material was considered extra purified. It was concentrated in an ultrafiltration centrifuge tube, and the resulting protein/lysozyme was used to determine its bactericidal potential against five bacterial strains, including three gram-positive (Bacillus subtilis 168, Micrococcus luteus, and Bacillus cereus) and two gram-negative (Salmonella typhimurium and Pseudomonas aeruginosa) strains. The results of ZOI and MIC/MBC showed that lysozyme had a higher antimicrobial activity against gram-positive than gram-negative bacterial strains. The results of the antibacterial activity of lysozyme were compared with those of ciprofloxacin (antibiotic). For this purpose, two indices were applied in the present study: antimicrobial index (AMI) and percent activity index (PAI). It was found that the purified lysozyme had a higher antibacterial activity against Bacillus cereus (AMI/PAI; 1.01/101) and Bacillus subtilis 168 (AMI/PAI; 1.03/103), compared to the antibiotic (ciprofloxacin) used in this study. Atomic force microscopy (AFM) was used to determine the bactericidal action of the lysozyme on the bacterial cell. The purified protein was further processed by gel column chromatography and the eluate was collected, its enzymatic activity was 21.93 U/mL, while the eluate was processed by native-PAGE. By this analysis, the un-denatured protein with enzymatic activity of 40.9 U/mL was obtained. This step shows that the protein (lysozyme) has an even higher enzymatic potential. To determine the specific peptides (in lysozyme) that may cause the bactericidal potential and cell lytic/enzymatic activity, the isolated protein (lysozyme) was further processed by the SDS-PAGE technique. SDS-PAGE analysis revealed different bands with sizes of 34 kDa, 24 kDa, and 10 kDa, respectively. To determine the chemical composition of the peptides, the bands (from SDS-PAGE) were cut, enzymatically digested, desalted, and analyzed by LC-MS (liquid chromatography-mass spectrometry). LC-MS analysis showed that the purified lysozyme had the following composition: the number of proteins in the sample was 56, the number of peptides was 124, and the number of PSMs (peptide spectrum matches) was 309. Among them, two peptides related to lysozyme and bactericidal activities were identified as: A0A1Q9G213 (N-acetylmuramoyl-L-alanine amidase) and A0A1Q9FRD3 (D-alanyl-D-alanine carboxypeptidase). The corresponding protein sequence and nucleic acid sequence were determined by comparison with the database.

Red-to-blue colorimetric probe based on biomass carbon dots for smartphone-integrated optosensing of Cu(II) and L-cysteine.

We constructed a smartphone-integrated optosensor with inexpensive, reversible, environmental friendly, and rapid adsorption to detect Cu(II) and L-cysteine (L-Cys). The key part of this study was to prepare a red-to-blue colorimetric probe from herbaceous andrographis paniculata using one-pot polymerization at room temperature. When Cu(II) existed, the red fluorescence on the surface of the core-shell probe was quenched, while the blue fluorescence of the core did not respond, because the colorimetric probe interacted with the Cu(II) on the surface of red CDs. After L-Cys added, it interacted with the Cu(II) to strip it from the surface of red CDs, resulting in the recovery of fluorescence response. Under optimal conditions, the detection limits of this method for Cu(II) and L-Cys were 71 nM and 12 nM, respectively. Further, the red-to-blue colorimetric probe was integrated into smartphone with a software application to convert fluorescent color images into specific red (R), green (G), and blue (B) values. The spiked recovery of Cu(II) and L-Cys in lake water was verified the feasibility of the developed optosensors with a recovery of 98.2-101.6 % and 103.3-121.6 %. This method for detecting Cu(II) and L-Cys can not only recognize metal ions from actual samples, but also effectively protect CDs from quenching and restore fluorescence.

Energy-efficient Preparation of Amino and Sulfhydryl Functionalized Biomass Carbon Dots via a Reverse Microemulsion for Specific Recognition of Fe3+ and L-cysteine.

Amino- and sulfhydryl- functionalized biomass carbon dots (BCDs) were prepared by one-pot reverse microemulsion for specific recognition of ferric ions (Fe3+) and L-cysteine (L-Cys). Green grapefruit peel was used as the carbon source while aminosilane and mercaptosilane were used as N- and S-supplier. Following the adsorption of Fe3+ on the surfaces of BCDs-NH2 and BCDs-SH, the fluorescence responses was quenched step by step, while adding L-Cys to the BCDs-NH2/Fe3+ system restored the fluorescence. The BCDs-NH2 and BCDs-SH system exhibited extremely low limits of detection for Fe3+ of 3.2 and 3.0 nM, respectively, within a wide linear ranges of 0.006-200 µM and 0.004-200 µM, respectively. The BCDs-NH2/Fe3+ systems were used as an optosensor for L-Cys in the concentration ranges of 0.08-30 and 30-1000 µM with a detection limit of 65 nM. Developed BCDs-NH2 and BCDs-SH were able to respond to Fe3+ in water samples with satisfactory recoveries of 100.1%-103.1% and 94.6%-108.5%, respectively, and the BCDs-NH2/Fe3+ system was also able to respond to BCDs-NH2/Fe3+ in actual lake water samples with recoveries from 87.3% to 98.8%. Meanwhile, The BCDs-NH2 exhibited good photoluminescence and stability, and the with a fluorescence quantum yield was as high as 25%. This work demonstrates the feasibility of using such materials to remove hazardous ions from water and employing the resulting complexes for optosensing in a sustainable manner.

Lysozyme and Its Application as Antibacterial Agent in Food Industry.

Lysozymes are hydrolytic enzymes characterized by their ability to cleave the ß-(1,4)-glycosidic bonds in peptidoglycan, a major structural component of the bacterial cell wall. This hydrolysis action compromises the integrity of the cell wall, causing the lysis of bacteria. For more than 80 years, its role of antibacterial defense in animals has been renowned, and it is also used as a preservative in foods and pharmaceuticals. In order to improve the antimicrobial efficacy of lysozyme, extensive research has been intended for its modifications. This manuscript reviews the natural antibiotic compound lysozyme with reference to its catalytic and non-catalytic mode of antibacterial action, lysozyme types, susceptibility and resistance of bacteria, modification of lysozyme molecules, and its applications in the food industry.

Biogenic Selenium Nanoparticles and Their Anticancer Effects Pertaining to Probiotic Bacteria-A Review.

Selenium nanoparticles (SeNPs) can be produced by biogenic, physical, and chemical processes. The physical and chemical processes have hazardous effects. However, biogenic synthesis (by microorganisms) is an eco-friendly and economical technique that is non-toxic to human and animal health. The mechanism for biogenic SeNPs from microorganisms is still not well understood. Over the past two decades, extensive research has been conducted on the nutritional and therapeutic applications of biogenic SeNPs. The research revealed that biogenic SeNPs are considered novel competitors in the pharmaceutical and food industries, as they have been shown to be virtually non-toxic when used in medical practice and as dietary supplements and release only trace amounts of Se ions when ingested. Various pathogenic and probiotic/nonpathogenic bacteria are used for the biogenic synthesis of SeNPs. However, in the case of biosynthesis by pathogenic bacteria, extraction and purification techniques are required for further useful applications of these biogenic SeNPs. This review focuses on the applications of SeNPs (derived from probiotic/nonpathogenic organisms) as promising anticancer agents. This review describes that SeNPs derived from probiotic/nonpathogenic organisms are considered safe for human consumption. These biogenic SeNPs reduce oxidative stress in the human body and have also been shown to be effective against breast, prostate, lung, liver, and colon cancers. This review provides helpful information on the safe use of biogenic SeNPs and their economic importance for dietary and therapeutic purposes, especially as anticancer agents.

Carbon Quantum Dots from Pomelo Peel as Fluorescence Probes for "Turn-Off-On" High-Sensitivity Detection of Fe3+ and L-Cysteine.

This study designed a "turn-off-on" fluorescence analysis method based on carbon quantum dots (CQDs) to detect metal ions and amino acids in real sample systems. CQDs were derived from green pomelo peel via a one-step hydrothermal process. The co-doped CQDs with N and S atoms imparted excellent optical properties (quantum yield = 17.31%). The prepared CQDs could be used as fluorescent "turn-off" probes to detect Fe3+ with a limit of detection of 0.086 µM, a linear detection range of 0.1-160 µM, and recovery of 83.47-106.53% in water samples. The quenched CQD fluorescence could be turned on after adding L-cysteine (L-Cys), which allowed detection of L-Cys with a detection limit of 0.34 µM and linear range of 0.4-85 µM. Recovery of L-Cys in amino acid beverage was 87.08-122.74%. Visual paper-based testing strips and cellulose/CQDs composite hydrogels could be also used to detect Fe3+ and L-Cys.

Biosynthesis of Selenium Nanoparticles (via Bacillus subtilis BSN313), and Their Isolation, Characterization, and Bioactivities.

Among the trace elements, selenium (Se) has great demand as a health supplement. Compared to its other forms, selenium nanoparticles have minor toxicity, superior reactivity, and excellent bioavailability. The present study was conducted to produce selenium nanoparticles (SeNPs) via a biosynthetic approach using probiotic Bacillus subtilis BSN313 in an economical and easy manner. The BSN313 exhibited a gradual increase in Se reduction and production of SeNPs up to 5-200 µg/mL of its environmental Se. However, the capability was decreased beyond that concentration. The capacity for extracellular SeNP production was evidenced by the emergence of red color, then confirmed by a microscopic approach. Produced SeNPs were purified, freeze-dried, and subsequently characterized systematically using UV-Vis spectroscopy, FTIR, Zetasizer, SEM-EDS, and TEM techniques. SEM-EDS analysis proved the presence of selenium as the foremost constituent of SeNPs. With an average particle size of 530 nm, SeNPs were shown to have a -26.9 (mV) zeta potential and -2.11 µm cm/Vs electrophoretic mobility in water. SeNPs produced during both the 24 and 48 h incubation periods showed good antioxidant activity in terms of DPPH and ABST scavenging action at a concentration of 150 µg/mL with no significant differences (p > 0.05). Moreover, 200 µg/mL of SeNPs showed antibacterial reactivity against Escherichia coli ATCC 8739, Staphylococcus aureus ATCC 9027, and Pseudomonas aeruginosa ATCC 25923. In the future, this work will be helpful to produce biogenic SeNPs using probiotic Bacillus subtilis BSN313 as biofactories, with the potential for safe use in biomedical and nutritional applications.

Self-Assembling Behavior of pH-Responsive Peptide A6K without End-Capping.

A short self-assembly peptide A6K (H2N-AAAAAAK-OH) with unmodified N- and C-terminus was designed, and the charge distribution model of this short peptide at different pH was established by computer simulation. The pH of the solution was adjusted according to the model and the corresponding self-assembled structure was observed using a transmission electron microscope (TEM). As the pH changes, the peptide will assemble into blocks or nanoribbons, which indicates that the A6K peptide is a pH-responsive peptide. Circular dichroism (CD) and molecular dynamics (MD) simulation showed that the block structure was formed by random coils, while the increase in ß-turn content contributes to the formation of intact nanoribbons. A reasonable explanation of the self-assembling structure was made according to the electrostatic distribution model and the effect of electrostatic interaction on self-assembly was investigated. This study laid the foundation for further design of nanomaterials based on pH-responsive peptides.

The relationship between patterns of insulin secretion and risks of gestational diabetes mellitus.

OBJECTIVE: To evaluate patterns of insulin secretion in pregnancy and analyze the association between insulin patterns and risks of gestational diabetes mellitus (GDM). METHODS: A prospective study was conducted to collect and analyze pregnant women's materials from January 2015 to December 2018. Pregnant women were grouped according to results of 75-g oral glucose tolerance test at 24-28 weeks of pregnancy: normal glucose tolerance (NGT) and GDM. Insulin secretion patterns were based on the time of peak(s) and shape of insulin secretion curve. The relationship between insulin secretion patterns and pregnant outcomes was analyzed. RESULTS: A total of 2432 pregnant women met the inclusion criteria during the study period. Among them, 737 (30.3%) women were grouped as GDM and 1695 (69.7%) as NGT. Type I insulin secretion represented the early phase of insulin secretion (peak time at 30 or 60 minutes), while type II represented the delayed peak of insulin secretion (peak time at 120 or 180 minutes). Logistic regression analysis showed that type II insulin secretion was a risk factor of pre-eclampsia, large-for-gestational-age, and neonatal hypoglycemia. CONCLUSION: The delayed insulin peak is a useful marker for risk of GDM and adverse pregnant outcomes in women with GDM.

Isolation of selenium-resistant bacteria and advancement under enrichment conditions for selected probiotic Bacillus subtilis (BSN313).

The main aim of this work was to screen, isolate, and identify a probiotic selenium (Se)-resistant strain of Bacillus subtilis, using the 16S rDNA sequencing approach and subsequently optimize conditions. Initially, conditions were enhanced in two univariate optimization environments: shakings flask and a bioreactor. After solving optimization for selected variables, conditions were further optimized using orthogonal array testing. The results were further evaluated by the analysis of variance, in support of Se enrichment. In a bioreactor, based on R and F values, the order of effect of selected conditions on Se enrichment was stirring speed > initial pH > temperature > Se addition time. The stirring speed of the bioreactor was most significant, due to the suspension of reduced Se, as it formed. After absolute optimization, strain BSN313 was able to enrich Se up to 2,123 µg/g of dry weight, which is 7.58 times greater than the baseline Se-resistance. PRACTICAL APPLICATIONS: Systematic studies of selenium enrichment conditions will facilitate the successful development of an organic selenium source and the safe use of Bacillus subtilis strain (BSN313) as a food supplement. Selenium-enriched probiotic bacteria are reported to provide many health benefits to the host, due to antipathogenic, antioxidative, anticarcinogenic, antimutagenic, and anti-inflammatory activities.

Production and Application of Biosurfactant Produced by Bacillus Licheniformis Ali5 in Enhanced Oil Recovery and Motor Oil Removal from Contaminated Sand.

The present study describes the production of biosurfactant from isolate B. licheniformis Ali5. Seven different, previously-reported minimal media were screened for biosurfactant production, and two selected media were further optimized for carbon source. Further, various fermentation conditions such as (pH 2-12, temperature 20-50 °C, agitation speed 100-300 rpm, NaCl (0-30 g·L-1) were investigated. The partially purified biosurfactant was characterized by Fourier transform infrared spectroscopy (FTIR) and matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF MS) and found a lipopeptide mixture, similar to lichenysin-A. Biosurfactant reduced surface tension from 72.0 to 26.21 ± 0.3 and interfacial tension by 0.26 ± 0.1 mN.m-1 respectively, biosurfactant yield under optimized conditions was 1 g·L-1, with critical micelle concentration (CMC) of 21 mg·L-1 with high emulsification activity of (E24) 66.4 ± 1.4% against crude oil. Biosurfactant was found to be stable over extreme conditions. It also altered the wettability of hydrophobic surface by changing the contact angle from 49.76° to 16.97°. Biosurfactant efficiently removed (70-79%) motor oil from sand, with an efficiency of more than 2 fold as compared without biosurfactant (36-38%). It gave 32% additional oil recovery over residual oil saturation upon application to a sand-packed column. These results are indicative of potential application of biosurfactant in wettability alteration and ex-situ microbial enhanced oil recovery.