Presence Of Riboflavin Research Articles

PH-Responsive Collagen Hydrogels Prepared by UV Irradiation in the Presence of Riboflavin

This study reveals the pH-responsive behavior of collagen hydrogels prepared using ultraviolet (UV) irradiation with riboflavin as a photosensitizer. By varying the UV exposure time, we modulated the crosslinking density, thereby influencing the mechanical properties and pH responsiveness. Rheological analysis confirmed successful network formation, whereas swelling studies revealed significant pH-dependent behavior, with maximum swelling at a pH of four and minimal swelling above a pH of six, demonstrating partial reversibility over multiple pH cycles. Mechanical testing showed a pH-dependent elastic modulus, which increased 10 fold from a pH of 6 to 10. Fibroblast proliferation assays confirmed the biocompatibility of the hydrogels, with cell growth positively correlating with the UV exposure time. This research demonstrates the potential of UV-crosslinked collagen hydrogels in biomedical applications, such as tissue engineering and drug delivery, where pH responsiveness is essential.

Riboflavin as a Coloring Agent of Tablets Affects the Photostability of Manidipine after the Change of Dosage Forms

Manidipine (MP) is widely used for reducing high blood pressure. Calslot® (CALS) tablets, which are the original MP medicines, and their generic medicines have been used for patients in clinical situations. The authors hypothesized that the photodegradability of MP drug substance in CALS tablets might be enhanced when the tablets were photo-exposed after the change of the dosage form by the presence of riboflavin (RF), which is utilized as a coloring agent and a well-known photosensitizer. The present study clarified that RF enhanced the photodegradation of MP when the powders and the suspensions of CALS tablets were ultraviolet light (UV) irradiated. The addition of RF to the suspension of MP standard substances also promoted MP photodegradation along with the increase of the generation rate of its main photoproduct, benzophenone. Finally, the authors performed the photostabilization of MP suspensions based on the addition of quercetin (QU), which is one of polyphenols and has both the antioxidative potency and the UV filtering potency. It is summarized that QU has a protective potency for MP’s own photodegradation, and it partially suppresses the photocatalytic effect of RF. Further studies focused on the photochemical behaviors of utilized additives for medicines are needed for their safe use.

Ultraviolet light and riboflavin accelerates red blood cell dysfunction in vitro and in a guinea pig transfusion model.

Quality assessment of modified or processed red blood cell (RBC) components, such as pathogen-reduced RBCs, using only in vitro testing may not always be predictive of in vivo performance. Mouse or rat in vivo models are limited by a lack of applicability to certain aspects of human RBC biology. Here, we used a guinea pig model to study the effects of riboflavin combined with UV light on the integrity of RBCs in vitro and following transfusion in vivo. Guinea pig RBCs were collected from whole blood (WB) treated with varying UV doses (10, 20, 40 or 80 J/mL) in the presence of riboflavin (UVR-RBCs). In vitro tests for UVR-RBCs included hemolysis, osmotic fragility, and cellular morphology by scanning electron microscopy. Guinea pigs transfused with one-day post-treatment UVR-RBCs were evaluated for plasma hemoglobin (Hb), non-transferrin bound iron (NTBI), total iron and Perls-detectable hemosiderin deposition in the spleen and kidney, and renal uptake of Hb. Acute RBC injury was dose dependently accelerated after treatment with UV light in the presence of riboflavin. Aberrant RBC morphology was evident at 20, 40, and 80 J/mL, and membrane lysis with Hb release was prominent at 80 J/mL. Guinea pigs transfused with 40 and 80 J/mL UVR-RBCs showed increased plasma Hb levels, and plasma NTBI was elevated in all UVR-RBC groups (10-80 J/mL). Total iron levels and Perls-hemosiderin staining in spleen and kidney as well as Hb uptake in renal proximal tubules were increased 8 hours post-transfusion with 40 and 80 J/mL UVR-RBCs. UVR-RBCs administered to guinea pigs increased markers of intravascular and extravascular hemolysis in a UV dose-dependent manner. This model may allow for the discrimination of RBC injury during testing of extensively processed RBCs intended for transfusion.

Electrochemically polymerized dopamine activated carbon paste sensor for selective and sensitive detection of rutin in the presence of riboflavin

In this work, a sensor was developed for the quantification of RT in food and medicine samples using electro-polymerization of dopamine (DA) on the surface of a carbon paste electrode in 0.2 M phosphate buffered saline (PBS) of 7.0 pH by cyclic voltammetry (CV). The surface properties of bare carbon paste electrode (BCPE) and poly (DA) modified carbon paste electrode (PDAMCPE) were studied using scanning electron microscopy (SEM), electrochemical impedance study (EIS), and CV techniques. Two electron two proton transfer process was seen during the electrochemical reaction of RT with adsorption-controlled process. Under optimum conditions, the peak current increased linearly with an increase in RT concentration for CV in the range of 0.4 to 2.0 μM and differential pulse voltammetry (DPV) in the range of 0.4 to 2.5 μM and the limit of detection (LOD) was found to be 0.0045 μΜ and for CV and 0.079 μM for DPV techniques respectively. Even in the presence of some interferents like organic compounds and metal ions, the developed electrode exhibits good anti-interference ability for RT, and good selectivity was obtained for RT in the presence of riboflavin (RF). The developed sensor presents good stability, repeatability, and reproducibility. The CV measurement was performed for the analysis of RT in real samples such as green tea and medicine samples which showed a good recovery and this work has been not reported previously.

Identification of β-aspartic semialdehyde and homocysteine as major reaction products of riboflavin-sensitized photooxidation of peptide-bound methionine

Methionine is an essential amino acid for mammals and it is limiting for monogastric animals. It can be oxidized easily by UV light. This could influence the bioaccessibility and bioavailability of methionine. In this work, the photosensitized degradation of peptide-bound methionine in the presence of riboflavin was investigated in a model system. Capillary electrophoresis was employed to analyze the time course of the degradation. The products were identified by liquid chromatography coupled to mass spectrometry (LC–MS/MS). Benzoyl methionine was degraded by 50% during UV irradiation in the presence of riboflavin after 5.0 min with 10 mol% riboflavin and 6.4 min with 3 mol% riboflavin. Homocysteine (16–20 mol%) and β-aspartic semialdehyde (ca. 30 mol%) were found as major degradation products next to methionine sulfoxide (ca. 25 mol%). A smaller molar ratio of riboflavin led to a higher formation of aspartic semialdehyde. The formation of homocysteine was paralleled by the formation of formaldehyde. Furthermore, the experiment was transferred to small peptides, which showed the analogous degradation products of peptide-bound methionine.

The combined effect of carbon starvation and exogenous riboflavin accelerated the Pseudomonas aeruginosa-induced nickel corrosion

The primary objective of this study is to elucidate the synergistic effect of an exogenous redox mediator and carbon starvation on the microbiologically influenced corrosion (MIC) of metal nickel (Ni) by nitrate reducing Pseudomonas aeruginosa. Carbon source (CS) starvation markedly accelerates Ni MIC by P. aeruginosa. Moreover, the addition of exogenous riboflavin significantly decreases the corrosion resistance of Ni. The MIC rate of Ni (based on corrosion loss volume) is ranked as: 10 % CS level + riboflavin > 100 % CS level + riboflavin > 10 % CS level > 100 % CS level. Notably, starved P. aeruginosa biofilm demonstrates greater aggressiveness in contributing to the initiation of surface pitting on Ni. Under CS deficiency (10 % CS level) in the presence of riboflavin, the deepest Ni pits reach a maximum depth of 11.2 μm, and the corrosion current density (icorr) peak at approximately 1.35 × 10−5 A·cm−2, representing a 2.6-fold increase compared to the full-strength media (5.25 × 10−6 A·cm−2). For the 10 % CS and 100 % CS media, the addition of exogenous riboflavin increases the Ni MIC rate by 3.5-fold and 2.9-fold, respectively. Riboflavin has been found to significantly accelerate corrosion, with its augmentation effect on Ni MIC increasing as the CS level decreases. Overall, riboflavin promotes electron transfer from Ni to P. aeruginosa, thus accelerating Ni MIC.

Photocrosslinked Co-Assembled Amino Acid Nanoparticles for Controlled Chemo/Photothermal Combined Anticancer Therapy.

Nanostructures formed from the self-assembly of amino acids are promising materials in many fields, especially for biomedical applications. However, their low stability resulting from the weak noncovalent interactions between the amino acid building blocks limits their use. In this work, nanoparticles co-assembled by fluorenylmethoxycarbonyl (Fmoc)-protected tyrosine (Fmoc-Tyr-OH) and tryptophan (Fmoc-Trp-OH) are crosslinked by ultraviolet (UV) light irradiation. Two methods are investigated to induce the dimerization of tyrosine, irradiating at 254nm or at 365nm in the presence of riboflavin as a photo-initiator. For the crosslinking performed at 254nm, both Fmoc-Tyr-OH and Fmoc-Trp-OH generate dimers. In contrast, only Fmoc-Tyr-OH participates in the riboflavin-mediated dimerization under irradiation at 365nm. The participation of both amino acids in forming the dimers leads to more stable crosslinked nanoparticles, allowing also to perform further chemical modifications for cancer applications. The anticancer drug doxorubicin (Dox) is adsorbed onto the crosslinked nanoparticles, subsequently coated by a tannic acid-iron complex, endowing the nanoparticles with glutathione-responsiveness and photothermal properties, allowing to control the release of Dox. A remarkable anticancer efficiency is obtained in vitro and in vivo in tumor-bearing mice thanks to the combined chemo- and photothermal treatment.

Cross-linked gelatin–riboflavin-based film incorporated with essential oils and silver nanoparticle by gamma-irradiation: A novel approach for extending the shelf life of meat

The gamma-irradiation is one of the most acceptable technologies to modify the biopolymer structure by forming a highly energetic electron. The effect of gamma-irradiation for cross-linking of gelatin (G) film in the presence of riboflavin (R) and the physicochemical properties of films were investigated. Different concentrations of R (0.3, 0.75, and 1.2% (w/w) based on the dry weight of G) and 3 doses of gamma-irradiation (5, 10, and 15 kGy) were used. The cross-linked film was incorporated with a mixture of essential oils (EOs) and silver nanoparticles (AgNPs) in order to use as an active packaging for prolonging the shelf life of meat. The results indicated 0.75% R (w/w) and 5 kGy irradiation had a positive effect on the tensile strength, water insolubility, and water barrier properties of the composite G film. Nevertheless, the reduction of film elongation (%), suggests the formation of compact structure and cross-linking through the generation of radicals. The infrared spectroscopy evaluation showed that proper irradiation dosages could produce cross-linking bonds in G film and form a denser network. In vitro and in situ microbial analyses showed that the active film incorporated with EO-AgNPs had antimicrobial activity against most spoilage and pathogenic bacteria and was able to prolong the shelf life of meat by up to 21 days. Therefore, irradiated G-R-EO-AgNP film with good mechanical and antimicrobial properties has the potential to be developed as a biodegradable food packaging material in the food industry.

Effect of external factors on the stability of sparkling wine quality during storage

The shelf life of sparkling wine is an indicator that can vary depending on the impact of a large number of external factors on the product. These factors include transportation conditions, storage temperature, humidity, exposure to light, and packaging quality. The current regulatory documentation regulates certain requirements for the storage conditions of sparkling wines, but they are often violated during transportation and storage, which leads to a significant deterioration in the quality of products. This review is devoted to the study of changes in the quality characteristics of wines during their storage and transportation. It is shown that the intensity of physicochemical changes occurring in wine during storage depends on temperature conditions, the presence of vibration and exposure to light. The issue of the influence of temperature storage conditions on the quality characteristics of wine products, including sparkling wines, has been studied in some detail. Most researchers are of the opinion that in order to maintain a stable quality of wine, it is necessary to store and transport it at a constant temperature not exceeding 20 °C. In scientific publications, there are different views on the problem of the influence of bottle color on the change in chemical composition and the preservation of the quality characteristics of both still and sparkling wines during storage. It has been found that bottles made of darker glass allow protecting wine to a greater extent from the effects of light, especially in its ultraviolet and visible short-wave (λ = 400 nm) range, and prevent the appearance of the so-called “taste of light” (gout de lumière) in the wine. It is shown that the causes of the appearance of the “taste of light” in champagne are sulfur-containing compounds formed as a result of photochemical reactions. It has been found that the intensity of these reactions depends on the degree of illumination, as well as the presence of riboflavin and iron (III) ions in the wine. It is concluded that controlling the amount of light to which wines are exposed can prevent deterioration of their quality during storage and extend the shelf life. The possibility of preventing photodegradation of wine through the use of various antioxidants is also considered. Based on the information available in scientific publications, the study of the development of negative changes in wine under the influence of light is an important aspect from the point of view of risk assessment during product storage.

Ultraviolet-induced mechanical augmentation of the degraded porcine aortic adventitia: Its significance for preventing aneurysmal rupture

In the previous studies, we have shown that the tunica adventitia of porcine abdominal aorta could be mechanically reinforced by irradiating it with ultraviolet A (UV-A) rays to promote the photocrosslinking of constitutive collagen, and we have proposed the method as a procedure to prevent or delay abdominal aortic aneurysm (AAA) ruptures. We have also demonstrated that UV irradiation-induced mechanical augmentation despite the degradation of the adventitia through collagenolysis in vitro. Considering that elastolysis is equally a relevant event in the pathogenesis of AAA, the degradation of elastin was also investigated in the present study. A total of 50 porcine aortas were used in the study. All processed samples were evaluated in a mechanical tester before and after degradation and/or irradiation. The adventitial layer was isolated and subjected to elastase for either 1 – 48 h. We found that both elastin and collagen were digested by elastase, with the former being completely digested after 48 h. The samples degraded for 1 h were subsequently irradiated with UV-A (365 nm) in the presence of riboflavin as a photoinitiator, a process that induced an enhancement of the strength and stiffness of the tissue. This is an indication that a partly degenerated aortic wall, like that in an aneurysmal region, can be reinforced mechanically by UV irradiation, possibly to the extent of delaying, or preventing altogether, the wall’s rupture. It appears important to irradiate the wall as early as possible.

Nitration of Tyrosine and Tyrosyl Residues in Myoglobin by the Action of Visible Light in the Presence of Riboflavin and Nitrite**

Nitration of Tyrosine and Tyrosyl Residues in Myoglobin by the Action of Visible Light in the Presence of Riboflavin and Nitrite**

Design, Synthesis, Characterization, and Evaluation of the Anti-HT-29 Colorectal Cell Line Activity of Novel 8-Oxyquinolinate-Platinum(II)-Loaded Nanostructured Lipid Carriers Targeted with Riboflavin.

Colorectal cancer is occasionally called colon or rectal cancer, depending on where cancer begins to form, and is the second leading cause of cancer death among both men and women. The platinum-based [PtCl(8-O-quinolinate)(dmso)] (8-QO-Pt) compound has demonstrated encouraging anticancer activity. Three different systems of 8-QO-Pt-encapsulated nanostructured lipid carriers (NLCs) with riboflavin (RFV) were investigated. NLCs of myristyl myristate were synthesized by ultrasonication in the presence of RFV. RFV-decorated nanoparticles displayed a spherical shape and a narrow size dispersion in the range of 144-175 nm mean particle diameter. The 8-QO-Pt-loaded formulations of NLC/RFV with more than 70% encapsulation efficiency showed sustained in vitro release for 24 h. Cytotoxicity, cell uptake, and apoptosis were evaluated in the HT-29 human colorectal adenocarcinoma cell line. The results revealed that 8-QO-Pt-loaded formulations of NLC/RFV showed higher cytotoxicity than the free 8-QO-Pt compound at 5.0 µM. All three systems exhibited different levels of cellular internalization. Moreover, the hemotoxicity assay showed the safety profile of the formulations (less than 3.7%). Taken together, RFV-targeted NLC systems for drug delivery have been investigated for the first time in our study and the results are promising for the future of chemotherapy in colon cancer treatment.

Single-step extraction of small-diameter single-walled carbon nanotubes in the presence of riboflavin.

We propose a novel approach to disperse and extract small-diameter single-walled carbon nanotubes (SWCNTs) using an aqueous solution of riboflavin and Sephacryl gel. The extraction of small-diameter semiconducting SWCNTs was observed, regardless of the initial diameter distribution of the SWCNTs. Dispersion of SWCNTs occurs due to the adsorption of π-conjugated isoalloxazine moieties on the surface of small-diameter nanotubes and interactions between hydroxy groups of ribityl chains with water. During the SWCNT extraction, specific adsorption of riboflavin to SWCNTs leads to the minimization of interactions between the SWCNTs and gel media. Our experimental findings are supported by ab initio calculations demonstrating the impact of the riboflavin wrapping pattern around the SWCNTs on their interaction with the allyl dextran gel.

Design, synthesis, and characterization of a novel dual cross-linked gelatin-based bioadhesive for hard and soft tissues adhesion capability

Many surgical treatments require a suitable tissue adhesive that maintains its performance in wet conditions and can be applied simultaneously for hard and soft tissues. In the present study, a dual cross-linked tissue adhesive was synthesized by mixing the gelatin methacryloyl (Gel-MA) and gelatin-dopamine conjugate (Gel-Dopa). The setting reaction was based on a photopolymerization process in the presence of a combination of riboflavin and triethanolamine and a chemical cross-linking process attributed to the genipin as a natural cross-linker. Modified gelatin macromolecules were characterized and the best wavelength for free radical generation in the presence of riboflavin was obtained. Tissue adhesives were prepared with 30% hydrogels of Gel-MA and Gel-Dopa with different ratios in distilled water. The gelation occurred in a short time after light irradiation. The chemical, mechanical, physical, and cytotoxicity properties of the tissue adhesives were evaluated. The results showed that despite photopolymerization, chemical crosslinking with genipin played a more critical role in the setting process. Water uptake, degradation behavior, cytotoxicity, and adhesion properties of the adhesives were correlated with the ratio of the components. The SEM images showed a porous structure that could ensure the entry of cells and nutrients into the surgical area. While acceptable properties in most experiments were observed, all features were improved as the Gel-Dopa ratio increased. Also, the obtained hydrogels revealed excellent adhesive properties, particularly with bone even after wet incubation, and it was attributed to the amount of gelatin-dopamine conjugate. From the obtained results, it was concluded that a dual adhesive hydrogel based on gelatin macromolecules could be a good candidate as a tissue adhesive in wet condition.

Deep blue autofluorescence reflects the oxidation state of human transthyretin

Human transthyretin (TTR) is a tetrameric protein transporting thyroid hormones and retinol. TTR is a neuroprotective factor and sensor of oxidative stress which stability is diminished due to mutations and aging, leading to amyloid deposition. Adverse environmental conditions, such as redox and metal ion imbalances, induce destabilization of the TTR structure. We have previously shown that the stability of TTR was disturbed by Ca2+ and other factors, including DTT, and led to the formation of an intrinsic fluorophore(s) emitting blue light, termed deep blue autofluorescence (dbAF). Here, we show that the redox state of TTR affects the formation dynamics and properties of dbAF. Free thiols lead to highly unstable subpopulations of TTR and the frequent ocurrence of dbAF. Oxidative conditions counteracted the destabilizing effects of free thiols to some extent. However, strong oxidative conditions led to modifications of TTR, which altered the stability of TTR and resulted in unique dbAF spectra. Riboflavin and/or riboflavin photoproducts bound to TTR and crosslinked TTR subunits. Riboflavin-sensitized photooxidation increased TTR unfolding, while photooxidation, either in the absence or presence of riboflavin, increased proteolysis and resulted in multiple oxidative modifications and dityrosine formation in TTR molecules. Therefore, oxidation can switch the role of TTR from a protective to pathogenic factor.

In Situ Crosslinkable Collagen-Based Hydrogels for 3D Printing of Dermis-Mimetic Constructs

Recently, several three-dimensional (3D) bioprinting techniques have emerged for the synthesis of 3D tissue analogs. Accordingly, many researchers have focused on the development of novel bioinks that can mimic the natural extracellular matrix with cytocompatibility and biofunctionality. Hyaluronic acid and collagen are the most abundant proteins in the extracellular matrix of the skin and are known to support several cellular behaviors. Herein, we developed tyramine-conjugated hyaluronic acid and collagen (HA-Tyr/Col-Tyr) hydrogel bioinks, which are photocrosslinkable in the presence of riboflavin and ammonium peroxydisulfate, to fabricate dermis-mimetic constructs. The physicochemical properties and 3D printability of the HA-Tyr/Col-Tyr hydrogel were examined. 3D printing of the lattice structure with the HA-Tyr/Col-Tyr hydrogel enabled a sophisticated micron-sized fine structure without any clogging or coagulation. Approximately 80% of normal human dermal fibroblasts (NHDFs) in the printed constructs were alive after 24 h of culture. Moreover, the 3D printed constructs supported 4.57-fold cell proliferation and 4.23-fold f-actin expansion over four days in culture, indicating that HA-Tyr/Col-Tyr hydrogels provide cytocompatible microenvironments. The findings of this study suggest that HA-Tyr/Col-Tyr hydrogels are promising candidates as bioinks for the 3D printing of dermis-mimetic constructs.

O7 - Synthesis and Incorporation of Quaternary Ammonium Silane Antimicrobial into Self-Crosslinked Type I Collagen Scaffold: A hybrid formulation for 3D Printing

To construct a novel 3D-biomaterial scaffold having a combination of a new quaternary ammonium silane (k21) antimicrobial impregnated in three-dimensional 3D collagen printing cross linked with Riboflavin in presence of d-alpha-tocopheryl poly(ethyleneglycol)-1000-succinate as a potential scaffold. Groups of 0.1 and 0.2% k21, and 0.1% and 0.2% CHX were prepared. k21/CHX with neutralized collagen was printed with BioX TM . Riboflavin was photo-activated and examined using epifluorescence for Aggregatibacter actinomycetemcomitans (7-days). Collagen was examined using TEM and measured for porosity, and shape-fitting. Raman and Tandem Mass/solid-state were performed with molecular-docking and circular dichroism. X-ray diffraction were conducted with rheological tests, contact angle and ninhydrin assay. k21 samples demonstrated collagen aggregates while 0.1%CHX and 0.2%CHX showed irregularities. Porosity of control and 0.1% and 0.2% k21 scaffolds showed no differences. Low contact angle, improved elastic-modulus, rigidity, and smaller strain in k21 groups were seen. Bacteria were reduced and strong organic intensities seen in k21 scaffolds. Simulation showed hydrophobicity and electrostatic interaction. Crosslinking was observed in 0.2%CHX/79%±9.9 and 0.2%k21/80%±12.3. Circular dichroism for k21 were suggestive of triple helix. XRD patterns appeared at d=5.97, 3.03, 2.78, 2.1 and 2.90 A o . 3D-printing of collagen impregnated with quaternary ammonium silane produced a promising scaffold with antimicrobial potency and structural stability.

Model Systems for Evidencing the Mediator Role of Riboflavin in the UVA Cross-Linking Treatment of Keratoconus.

Riboflavin under UVA radiation generates reactive oxygen species (ROS) that can induce various changes in biological systems. Under controlled conditions, these processes can be used in some treatments for ocular or dermal diseases. For instance, corneal cross-linking (CXL) treatment of keratoconus involves UVA irradiation combined with riboflavin aiming to induce the formation of new collagen fibrils in cornea. To reduce the damaging effect of ROS formed in the presence of riboflavin and UVA, the CXL treatment is performed with the addition of polysaccharides (dextran). Hyaluronic acid is a polysaccharide that can be found in the aqueous layer of the tear film. In many cases, keratoconus patients also present dry eye syndrome that can be reduced by the application of topical solutions containing hyaluronic acid. This study presents physico-chemical evidence on the effect of riboflavin on collagen fibril formation revealed by the following methods: differential scanning microcalorimetry, rheology, and STEM images. The collagen used was extracted from calf skin that contains type I collagen similar to that found in the eye. Spin trapping experiments on collagen/hyaluronic acid/riboflavin solutions evidenced the formation of ROS species by electron paramagnetic resonance measurements.

Synthesis and Incorporation of Quaternary Ammonium Silane Antimicrobial into Self-Crosslinked Type I Collagen Scaffold: A Hybrid Formulation for 3D Printing.

Novel 3D-biomaterial scaffold is constructed having a combination of a new quaternary ammonium silane (k21) antimicrobial impregnated in 3D collagen printed scaffolds cross linked with Riboflavin in presence of d-alpha-tocopheryl poly(ethyleneglycol)-1000-succinate. Groups of "0.1% and 0.2% k21", and "0.1% and 0.2% Chlorhexidine (CHX)" are prepared. k21/CHX with neutralized collagen is printed with BioX. Riboflavin is photo-activated and examined using epifluorescence for Aggregatibacter actinomycetemcomitans (7-days). Collagen is examined using TEM and measured for porosity, and shape-fitting. Raman and tandem mass/solid-state are performed with molecular-docking and circular-dichroism. X-ray diffractions, rheological tests, contact angle, and ninhydrin assay are conducted. k21 samples demonstrated collagen aggregates while 0.1% CHX and 0.2% CHX showed irregularities. Porosity of control and "0.1% and 0.2% k21" scaffolds show no differences. Low contact angle, improved elastic-modulus, rigidity, and smaller strain in k21 groups are seen. Bacteria are reduced and strong organic intensities are seen in k21 scaffolds. Simulation shows hydrophobicity/electrostatic interaction. Crosslinking is observed in 0.2% CHX/79% and 0.2% k21/80%. Circular dichroism for k21 are suggestive of triple helix. XRD patterns appear at d = 5.97, 3.03, 2.78, 2.1, and 2.90 A°. 3D-printing of collagen impregnated with quaternary ammonium silane produces a promising scaffold with antimicrobial potency and structural stability.

Metarhizium robertsii protease and conidia production, response to heat stress and virulence against Aedes aegypti larvae

Nutritional factors exert significant influence on the growth of entomopathogenic fungi, one of the main agents employed commercially in the biological control of arthropods. Thus, the objective of this work is to optimize the culture medium and solid fermentation time for production of proteases and conidia of Metarhizium robertsii ARSEF 2575 and to evaluate the interference of riboflavin and salts on virulence and resistance to abiotic stress factors. In the first step, nine groups were separated: negative control, positive control, and seven supplementation groups: ammonium nitrate, ammonium chloride, potassium nitrate, sodium nitrate, ammonium sulfate, ammonium phosphate, urea. Sodium nitrate showed significant difference in protease production at the time of 20 days of solid fermentation. Then, different concentrations of sodium nitrate and riboflavin as supplement were evaluated. Response surface methodology demonstrated that riboflavin and sodium nitrate influence proteolytic activity and conidia production, but without synergism. Supplementation of the medium with the optimal concentration of sodium nitrate and riboflavin did not interfere with the germination of conidia without exposure to abiotic stress, but did increase the thermotolerance of conidia. The presence of riboflavin and sodium nitrate at optimal concentrations in the culture medium did not alter fungal virulence with and without exposure to heat stress, varying according to the presence or absence of the supernatant during exposure, evidencing that resistance to heat exposure is multifactorial and dependent on intra- and extracellular factors. Moreover, the supplementation increased the larvicidal activity of the supernatant against Aedes aegypti.