Investigating the Anti-Inflammatory Properties and Skin Penetration Ability of Cornelian Cherry (Cornus mas L.) Extracts.

Plant extracts can be a valuable source of biologically active compounds in many cosmetic preparations. Their effect depends on the phytochemicals they contain and their ability to penetrate the skin. Therefore, in this study, the possibility of skin penetration by phenolic acids contained in dogwood extracts of different fruit colors (yellow, red, and dark ruby red) prepared using different extractants was investigated. These analyses were performed using a Franz chamber and HPLC-UV chromatography. Moreover, the antioxidant properties of the tested extracts were compared and their impact on the intracellular level of free radicals in skin cells was assessed. The cytotoxicity of these extracts towards keratinocytes and fibroblasts was also analyzed and their anti-inflammatory properties were assessed using the enzyme-linked immunosorbent assay (ELISA). The analyses showed differences in the penetration of individual phenolic acids into the skin and different biological activities of the tested extracts. None of the extracts had cytotoxic effects on skin cells in vitro, and the strongest antioxidant and anti-inflammatory properties were found in dogwood extracts with dark ruby red fruits.

Phenolic compound profiling and antioxidant potential of different types of Schisandra henryi in vitro cultures.

Schisandra henryi is an endemic species of medicinal potential known from traditional Chinese medicine. As part of this study, a complex biotechnological and phytochemical assessment was conducted on S. henryi with a focus on phenolic compounds and antioxidant profiling. The following in vitro cultures were tested: microshoot agar and callus, microshoot agitated, and suspension, along with the microshoot culture in PlantForm bioreactors. Qualitative profiling was performed by ultra-high-performance liquid chromatography with a photodiode array detector coupled with ion-trap mass spectrophotometry with electrospray ionization and then quantitative analysis by high-performance liquid chromatography with a diode array detector using standards. In the extracts, mainly the compounds from procyanidins were identified as well as phenolic acids (neochlorogenic acid, caffeic acid, protocatechuic acid) and catechin. The highest content of phenolic compounds was found for in vitro agar microshoot culture (max. total content 229.87 mg/100 g DW) and agitated culture (max. total content 22.82 mg/100 g DW). The max. TPC measured using the Folin-Ciocalteu assay was equal to 1240.51 mg GAE/100 g DW (agar microshoot culture). The extracts were evaluated for their antioxidant potential by the DPPH, FRAP, and chelate iron ion assays. The highest potential was indicated for agar microshoot culture (90% of inhibition and 59.31 nM/L TEAC, respectively). The research conducted on the polyphenol profiling and antioxidant potential of S. henryi in vitro culture extracts indicates the high therapeutic potential of this species. KEY POINTS: • Different types of S. henryi in vitro cultures were compared for the first time. • The S. henryi in vitro culture strong antioxidant potential was determined for the first time. • The polyphenol profiling of different types of S. henryi in vitro cultures was shown.

Green and Efficient Extraction of Phenolic Components from Plants with Supramolecular Solvents: Experimental and Theoretical Studies.

The supramolecular solvent (SUPRAS) has garnered significant attention as an innovative, efficient, and environmentally friendly solvent for the effective extraction and separation of bioactive compounds from natural resources. However, research on the use of a SUPRAS for the extraction of phenolic compounds from plants, which are highly valued in food products due to their exceptional antioxidant properties, remains scarce. The present study developed a green, ultra-sound-assisted SUPRAS method for the simultaneous determination of three phenolic acids in Prunella vulgaris using high-performance liquid chromatography (HPLC). The experimental parameters were meticulously optimized. The efficiency and antioxidant properties of the phenolic compounds obtained using different extraction methods were also compared. Under optimal conditions, the extraction efficiency of the SUPRAS, prepared with octanoic acid reverse micelles dispersed in ethanol-water, significantly exceeded that of conventional organic solvents. Moreover, the SUPRAS method demonstrated greater antioxidant capacity. Confocal laser scanning microscopy (CLSM) images revealed the spherical droplet structure of the SUPRAS, characterized by a well-defined circular fluorescence position, which coincided with the position of the phenolic acids. The phenolic acids were encapsulated within the SUPRAS droplets, indicating their efficient extraction capacity. Furthermore, molecular dynamics simulations combined with CLSM supported the proposed method's mechanism and theoretically demonstrated the superior extraction performance of the SUPRAS. In contrast to conventional methods, the higher extraction efficiency of the SUPRAS can be attributed to the larger solvent contact surface area, the formation of more types of hydrogen bonds between the extractants and the supramolecular solvents, and stronger, more stable interaction forces. The results of the theoretical studies corroborate the experimental outcomes.

Fabrication of multifunctional facial masks from phenolic acid grafted chitosan/collagen peptides via aqueous electrospinning.

Facial masks have become ubiquitous in our daily life to endow skin enough moisture and activated nutrition through mask nonwovens infused with skincare ingredients. However, the active nutrients in wet masks are prone to deterioration and deactivation. Herein, a novel multifunctional nanofiber dry mask was successfully prepared using aqueous-electrospun phenolic acid grafted chitosan/collagen peptides. When used, the functional nanofibers in the mask dissolve through spraying moisture, activating active ingredients in response to water and providing in-situ free radical scavenging, moisturizing and antibacterial effects to the skin. In this work, a series of gallic acid (GA), caffeic acid (CA), and protocatechuic acid (PA) have been studied to be grafted with chitosan to improve water solubility of chitosan (CS). Also, through aqueous electrospinning of phenolic acid-grafted chitosan/collagen peptides, a one-step green multifunctional nanofiber mask was obtained. The results showed that the mask had a 12.14 % moisturizing rate and a 94.09 % activity for removing free radicals from the skin after encountering moisture. Considering its high efficiency, controllable function release, and easy processability, the nanofiber multifunctional mask may provide a competitive alternative to facial masks and promote potential value-added applications of bio-based macro-molecules.

Chemical Profile and Biological Potential of Scaptotrigona Bee Products (Hymenoptera, Apidae, Meliponini): An Review.

Stingless bees belong to the Meliponini tribe and are widely distributed in the tropics and subtropics, where they perform important ecological services. Among the best distributed groups of stingless bees is the genus Scaptotrigona, which includes 22 species distributed throughout the neotropical region, including the area from Mexico to Argentina. Bees of this genus are responsible for the production of products such as honey, propolis, geopropolis and fermented pollen ("saburá"). This review aimed to provide an overview of the chemical composition and biological activities associated with derived products from stingless bees of the genus Scaptotrigona. The bibliographic review was carried out through searches in the Scopus, Web of Science, ScienceDirect and PubMed databases, including publications from 2003 to January 2023. The study of the chemodiversity of products derived from Scaptotrigona demonstrated the mainly presence of flavonoids, phenolic acids, terpenoids and alkaloids. It was also demonstrated that products derived from bees of the genus Scaptotrigona exhibit a wide range of biological effects, such as antibacterial, antioxidant, anti-inflammatory and antifungal activities, among other bioactivities. This review provides an overview of phytochemical and pharmacological investigations of the genus Scaptotrigona. However, it is essential to clarify the toxicity and food safety of these products.

Modified chitosan with different phenolic acids: Characterization, physicochemical properties, and biological activity.

This study synthesized five phenolic acid-chitosan copolymers utilizing the carbodiimide-mediated chemical crosslinking reaction. Comprehensive evaluations were conducted on their structural attributes, physicochemical properties, and biological activities. Fourier transform infrared confirmed successful grafting of phenolic acids onto chitosan via amide linkages. Additionally, ultraviolet-visible absorption spectroscopy and proton nuclear magnetic resonance analyses revealed novel absorption peaks between 200 and 400 nm and 6.0-8.0 ppm, respectively, attributable to the incorporated phenolic acids. Notably, the chitosan-gentisate acid copolymer exhibited significantly enhanced biological activity (p < 0.05) compared to pure chitosan and the other four conjugates, attributed to its highest grafting degree of approximately 295.93 mg/g. These modified chitosan derivatives effectively preserved the quality of sea bass (Lateolabrax japonicus) during refrigerated storage, extending its shelf-life by up to 9 days, 7 days, and 4 days relative to control, chitosan, and gentisate acid groups.

Synthesis and its biological activity of carboxymethyl hemicellulose p-hydroxybenzoate (P-CMHC).

Hemicellulose extracted from ecalyptus APMP pulping waste liquor and undergoes etherification modification to produce carboxymethyl hemicellulose (CMHC). Subsequently, CMHC undergoes esterification reaction with p-hydroxybenzoic acid to synthesize a novel polysaccharide-based preservative known as carboxymethyl hemicellulose p-hydroxybenzoate (P-CMHC). The synthesis conditions of P-CMHC were optimized using the response surface methodology, resulting in an optimal esterification condition that achieved a degree of substitution of 0.232. P-CMHC exhibits excellent antioxidant activity, including 2,2-diphenyl-1-picrylhydrazyl (DPPH) and hydroxyl radical scavenging activities. Additionally, it demonstrates favorable hygroscopic and moisturizing properties. Thiazole blue (MTT) experiments evaluating cell proliferation rate indicate that P-CMHC possesses negligible cytotoxicity, making it a promising, safe, and healthy preservative. Consequently, it can be considered as a new material for applications in the fields of biomedicine, food, and cosmetics.

Effects of grafting methods and raw materials on the physicochemical properties and biological activities of phenolic acids grafted oat ß-glucan.

Phenolic acids are commonly used as food biological preservatives. Grafting phenolic acids onto polysaccharides could effectively enhance their biological activities and environmental stability to varying degrees. However, grafting methods and raw materials could affect the physical properties and biological activities of the phenolic acid-grafted polysaccharides. In this study, caffeic acid (CA) and gallic acid (GA) were grafted onto oat ß-glucan (OG) and hydrolyzed oat ß-glucan (OGH) through N,N'-carbonyldiimidazole-mediated (CDI) and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride coupling N-hydroxysuccinimide (EDC/NHS) methods. Graft modification decreased the crystallinity and thermal stability of the conjugates, but retained good bioactivities for the conjugates. The antioxidant and bacteriostatic activities of the conjugates prepared by the EDC method were better than those of the CDI method, and the OGH-conjugates showed better biological activities than OG-conjugates. EDC-GAOGH showed best DPPH (89.78%) and ABTS (92.32%) scavenging activities. The inhibitory effect of EDC-GAOGH on Escherichia coli was significantly better than that of EDC-CAOGH, but for Staphylococcus aureus, the results are opposite, which indicating that different phenolic acid grafting products have different inhibitory effects on pathogenic microbes. In general, grafting phenolic acids onto OGH using EDC method is an effective strategy for preparing food biological preservative.

Effect of co-pigments on anthocyanins of Rhododendron arboreum and insights into interaction mechanism.

The impact of intermolecular copigmentation between five phenolic acids, two flavonoid and three amino acids with R. arboreum anthocyanins (ANS) and its isolated cyanidin-3-O-monoglycosides were investigated through experimental and theoretical approach. On addition of different copigments, phenolic acid induced strong hyperchromic (0.26-0.55 nm) and bathochromic shift (6.6-14.2 nm). The color intensity and stability of ANS with, storage at 4 °C & 25 °C, sunlight, oxidation and heat were evaluated by chromaticity, anthocyanin content, kinetic and structural simulation analysis. The strongest copigmentation reaction was observed with narningin (NA) and also showed high thermostability and highest half-life i.e. 3.39 h-1.24 h at 90-160 °C. The cyanidin-3-O-monoglycosides were analysed for their copigmentation effect and observations revealed that NA displayed best copigmentation effect to cyanidin-3-O-arabinoside (B) followed by cyanidin-3-O-galactoside (A), and cyanidin-3-O-rhamnoside (C). Additionally, structural simulation and steered molecular dynamics insights NA is the most favourable co-pigment involving π-π stacking and H-bonding.

Megalochelin, a Tridecapeptide Siderophore from a Talented Streptomycete.

Streptomycetes are bacteria known for their extraordinary biosynthetic capabilities. Herein, we describe the genome and metabolome of a particularly talented strain, Streptomyces ID71268. Its 8.4-Mbp genome harbors 32 bioinformatically predicted biosynthetic gene clusters (BGCs), out of which 10 are expressed under a single experimental condition. In addition to five families of known metabolites with previously assigned BGCs (nigericin, azalomycin F, ectoine, SF2766, and piericidin), we were able to predict BGCs for three additional metabolites: streptochlorin, serpetene, and marinomycin. The strain also produced two families of presumably novel metabolites, one of which was associated with growth inhibitory activity against the human opportunistic pathogen Acinetobacter baumannii in an iron-dependent manner. Bioassay-guided fractionation, followed by extensive liquid chromatography-mass spectrometry (LC-MS) and NMR analyses, established that the molecule responsible for the observed antibacterial activity is an unusual tridecapeptide siderophore with a ring-and-tail structure: the heptapeptide ring is formed through a C-C bond between a 2,3-dihydroxybenzoate (DHB) cap on Gly1 and the imidazole moiety of His7, while the hexapeptide tail is sufficient for binding iron. This molecule, named megalochelin, is the largest known siderophore. The megalochelin BGC encodes a 13-module nonribosomal peptide synthetase for the synthesis of the tridecapeptide, and a copper-dependent oxidase, likely responsible for the DHB-imidazole cross-link, whereas the genes for synthesis of the DHB starter unit are apparently specified in trans by a different BGC. Our results suggest that prolific producers of specialized metabolites may conceal hidden treasures within a background of known compounds.

Reply to Zarguan et al. Comment on "Budryn et al. Hydroxybenzoic Acids as Acetylcholinesterase Inhibitors: Calorimetric and Docking Simulation Studies. Nutrients 2022, 14, 2476".

We thank Prof. Zarguan's group for their comment [...].

In Vitro Reconstitution of Fimsbactin Biosynthesis from Acinetobacter baumannii.

Siderophores produced via nonribosomal peptide synthetase (NRPS) pathways serve as critical virulence factors for many pathogenic bacteria. Improved knowledge of siderophore biosynthesis guides the development of inhibitors, vaccines, and other therapeutic strategies. Fimsbactin A is a mixed ligand siderophore derived from human pathogenic Acinetobacter baumannii that contains phenolate-oxazoline, catechol, and hydroxamate metal chelating groups branching from a central l-Ser tetrahedral unit via amide and ester linkages. Fimsbactin A is derived from two molecules of l-Ser, two molecules of 2,3-dihydroxybenzoic acid (DHB), and one molecule of l-Orn and is a product of the fbs biosynthetic operon. Here, we report the complete in vitro reconstitution of fimsbactin A biosynthesis in a cell-free system using purified enzymes. We demonstrate the conversion of l-Orn to N1-acetyl-N1-hydroxy-putrescine (ahPutr) via ordered action of FbsJ (decarboxylase), FbsI (flavin N-monooxygenase), and FbsK (N-acetyltransferase). We achieve conversion of l-Ser, DHB, and l-Orn to fimsbactin A using FbsIJK in combination with the NRPS modules FbsEFGH. We also demonstrate chemoenzymatic conversion of synthetic ahPutr to fimsbactin A using FbsEFGH and establish the substrate selectivity for the NRPS adenylation domains in FbsH (DHB) and FbsF (l-Ser). We assign a role for the type II thioesterase FbsM in producing the shunt metabolite 2-(2,3-dihydroxyphenyl)-4,5-dihydrooxazole-4-carboxylic acid (DHB-oxa) via cleavage of the corresponding thioester intermediate that is tethered to NRPS peptidyl carrier domains during biosynthetic assembly. We propose a mechanism for branching NRPS-derived peptides via amide and ester linkages via the dynamic equilibration of N-DHB-Ser and O-DHB-Ser thioester intermediates via hydrolysis of DHB-oxa thioester intermediates. We also propose a genetic signature for NRPS "branching" in the presence of a terminating C-T-C motif (FbsG).

Synthesis, Characterization, and the Antioxidant Activity of Phenolic Acid Chitooligosaccharide Derivatives.

A series of phenolic acid chitooligosaccharide (COS) derivatives synthesized by two mild and green methods were illuminated in this paper. Seven phenolic acids were selected to combine two kinds of COS derivatives: the phenolic acid chitooligosaccharide salt derivatives and the phenolic-acid-acylated chitooligosaccharide derivatives. The structures of the derivatives were characterized by FT-IR and 1H NMR spectra. The antioxidant experiment results in vitro (including DPPH-radical scavenging activity, superoxide-radical scavenging activity, hydroxyl-radical scavenging ability, and reducing power) demonstrated that the derivatives exhibited significantly enhanced antioxidant activity compared to COS. Moreover, the study showed that the phenolic acid chitooligosaccharide salts had stronger antioxidant activity than phenolic-acid-acylated chitooligosaccharide. The cytotoxicity assay of L929 cells in vitro indicated that the derivatives had low cytotoxicity and good biocompatibility. In conclusion, this study provides a possible synthetic method for developing novel and nontoxic antioxidant agents which can be used in the food and cosmetics industry.

Rare ß-Resorcylic Acid Derivatives from a Halophyte-Associated Fungus Colletotrichum gloeosporioides JS0419 and Their Antifungal Activities.

Six new ß-resorcylic acid derivatives (1-5 and 7) were isolated from a halophyte-associated fungus, Colletotrichum gloeosporioides JS0419, together with four previously reported ß-resorcylic acid lactones (RALs). The relative and absolute stereochemistry of 1 was completely established by a combination of spectroscopic data and chemical reactions. The structures of the isolated compounds were elucidated by analysis of HRMS and NMR data. Notably, compounds 1-3 had a ß-resorcylic acid harboring a long unesterified aliphatic side chain, whereas the long aliphatic chains were esterified to form macrolactones in 4-9. Among the isolated compounds, monocillin I and radicicol showed potent antifungal activities against Cryptococcus neoformans, comparable to clinically available antifungal agents and radicicol showed weak antifungal activity against Candida albicans. These findings provide insight into the chemical diversity of fungal RAL-type compounds and their pharmacological potential.

The Antioxidant, Cytotoxic and Antimicrobial Potential of Phenolic Acids-Enriched Extract of Elicited Hairy Roots of Salvia bulleyana.

Hairy root cultures are valuable sources of a range of phytochemicals. Among them, Salvia bulleyana root culture is a promising source of polyphenols, especially rosmarinic acid (RA), a phenolic acid depside with pleiotropic activity and a wide application in medicine and cosmetology. The aim of the study was to enhance the culture productivity by finding suitable elicitation protocol and to determine its biological potential in terms of antioxidant, anticancer and antimicrobial properties. The total content of phenols and the levels of particular constituents in root extracts were analyzed using HPLC-PDA. Among four elicitors tested (yeast extract; methyl jasmonate, MJA; trans-anethol; and cadmium chloride), MJA was found to be the most effective. The greatest boost in phenolic production (up to 124.4 mg/g dry weight) was observed after three-day treatment with MJA at 100 µM, with an almost 100% improvement compared to the controls (non-treated root culture). The hydromethanolic extract from the elicited culture exhibited strong antioxidant activity with IC50 values of 11.1 µg/mL, 6.5 µg/mL and 69.5 µg/mL for DPPH (2,2-diphenyl-1-picrylhydrazyl), ABTS (2,2-azinobis-(3-ethylbenzthiazoline-6-sulfonic acid)) and superoxide anion radical, respectively. Moreover, in concentrations of 0.5-5 mg/mL the extract inhibited the growth of LoVo, AGS and HeLa cell lines, but was safe for the L929 cells up to the concentration of 5 mg/mL. The extract also exhibited moderate antimicrobial activity. Thus, the results confirmed that elicitation can be a beneficial strategy for increase the phenolic acid biosynthesis in hairy roots of S. bulleyana, and that such a highly productive culture can show significant biological potential.

Determination of Phenolic Compounds in Beverages by Three-Flow Channel Isocratic HPLC with Electrochemical Detections Using a Column-Switching Technique.

For the quantitative analysis of phenolic compounds in beverage samples, a three-flow channel isocratic HPLC with electrochemical detection (3LC-ECD) system was devised using a column-switching technique. Phenolic compounds with significantly different hydrophobicity (the range of calculated log P: -0.77 to 3.02) were simultaneously measured to draw three chromatograms by the 3LC-ECD; the peaks of gallic acid (GA), protocatechuic acid (PCA), and gallocatechin (GC) were observed at electrochemical detector 1 (D1) within 42 min, the peaks of procyanidin B3 (B3), epigallocatechin (EGC), catechin (C), epicatechin (EC), procyanidin B2 (B2), ethyl gallate (Eg), and epigallocatechin gallate (EGCg) were observed at D2 within 50 min, and the peaks of epicatechin gallate (ECg), gallocatechin gallate (GCg), catechin gallate (Cg), propyl gallate (Pg), and resveratrol (RVT) were observed at D3 within 70 min. The relationships between the phenolic compound concentrations and their chromatographic peak heights gave good linearity with correlation coefficients of more than 0.998. The detection limits of the phenolic compounds by the 3LC-ECD ranged from 0.6 to 3.0 µg/L. Further, the phenolic compound concentrations of commercially available teas and wines were determined with a relative standard deviation (RSD) of less than 4.9% (n = 6), and their recoveries ranged from 91 to 109%. These results indicate that the 3LC-ECD system provided an accurate, precise, and specific determination of the phenolic compounds in beverages without affecting the matrices derived from these samples.

Identification of key phenolic compounds responsible for antioxidant activities of free and bound fractions of blackberry varieties' extracts by boosted regression trees.

BACKGROUND: Free fractions of different blackberry varieties' extracts are high in phenolic compounds with antioxidant activities. However, the phenolic profiles and antioxidant activities against peroxyl radicals of bound fractions of different blackberry varieties' extracts have not been previously reported. In addition, what the key antioxidant phenolic compounds are in free and bound fractions of blackberry extracts remain unknown. This study aimed to investigate the phenolic profiles and antioxidant activities of free and bound fractions of eight blackberry varieties' extracts and reveal the key antioxidant phenolic compounds by boosted regression trees. RESULTS: Fifteen phenolics (three anthocyanins, four flavonols, three phenolic acids, two proanthocyanidins, and three ellagitannins) were identified in blackberry by ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry. Ferulic acid, ellagic acid, procyanidin C1, kaempferol-O-hexoside, ellagitannins hex, and gallic acid were major bound phenolics. Bound fractions of eight blackberry varieties' extracts were high in phenolics and showed great antioxidant activity. Boosted regression trees analysis showed that cyanidin-3-O-glucoside and chlorogenic acid were the most significant compounds, contributing 48.4% and 15.9% respectively to the antioxidant activity of free fraction. Ferulic acid was the most significant antioxidant compound in bound fraction, with a contribution of 61.5%. Principal component analysis showed that Kiowa was the best among the eight varieties due to its phenolic profile and antioxidant activity. CONCLUSION: It was concluded that blackberry varieties contained high amounts of bound phenolics, which confer health benefits through reducing oxidative stress. Ferulic acid was the key compound to explain the antioxidant activities of bound fractions. © 2021 Society of Chemical Industry.

Working with the natural complexity: Selection and characterization of black cohosh root extract for use in toxicology testing.

Black cohosh (Actaea racemosa L.) is a botanical supplement marketed to women of all ages. Due to paucity of data to assess the safe use, the National Toxicology Program (NTP) is evaluating the toxicity of black cohosh. The use of an authentic, quality material is imperative to generate robust data. Because botanical materials are complex mixtures with variable composition, the selection of a material is challenging. We describe selection and phytochemical characterization of an unformulated black cohosh root extract (i.e., an extract that serves as source material for a formulated product) to be used in the NTP assessments. A material was selected using a combination of non-targeted and targeted chemical analyses, including confirmation of authenticity, absence of contaminants and adulterants, and similarity to a popular black cohosh product used by consumers. Thirty-nine constituents covering three major classes, triterpene glycosides, phenolic acids, and alkaloids were identified. Among constituents quantified, triterpene glycosides made up approximately 4.7% (w/w) with total constituents quantified making up 5.8% (w/w) of the extract. Non-targeted chemical analysis followed by chemometric analysis of various materials sold as black cohosh, and reference materials for black cohosh and other Actaea species further confirmed the suitability of the selected extract for use.

Determination of the Protein-Protein Interactions within Acyl Carrier Protein (MmcB)-Dependent Modifications in the Biosynthesis of Mitomycin.

Mitomycin has a unique chemical structure and contains densely assembled functionalities with extraordinary antitumor activity. The previously proposed mitomycin C biosynthetic pathway has caused great attention to decipher the enzymatic mechanisms for assembling the pharmaceutically unprecedented chemical scaffold. Herein, we focused on the determination of acyl carrier protein (ACP)-dependent modification steps and identification of the protein-protein interactions between MmcB (ACP) with the partners in the early-stage biosynthesis of mitomycin C. Based on the initial genetic manipulation consisting of gene disruption and complementation experiments, genes mitE, mmcB, mitB, and mitF were identified as the essential functional genes in the mitomycin C biosynthesis, respectively. Further integration of biochemical analysis elucidated that MitE catalyzed CoA ligation of 3-amino-5-hydroxy-bezonic acid (AHBA), MmcB-tethered AHBA triggered the biosynthesis of mitomycin C, and both MitB and MitF were MmcB-dependent tailoring enzymes involved in the assembly of mitosane. Aiming at understanding the poorly characterized protein-protein interactions, the in vitro pull-down assay was carried out by monitoring MmcB individually with MitB and MitF. The observed results displayed the clear interactions between MmcB and MitB and MitF. The surface plasmon resonance (SPR) biosensor analysis further confirmed the protein-protein interactions of MmcB with MitB and MitF, respectively. Taken together, the current genetic and biochemical analysis will facilitate the investigations of the unusual enzymatic mechanisms for the structurally unique compound assembly and inspire attempts to modify the chemical scaffold of mitomycin family antibiotics.

Preclinical Pharmacokinetics and Acute Toxicity in Rats of 5-{[(2E)-3-Bromo-3-carboxyprop-2-enoyl]amino}-2-hydroxybenzoic Acid: A Novel 5-Aminosalicylic Acid Derivative with Potent Anti-Inflammatory Activity.

Compound 5-{[(2E)-3-bromo-3-carboxyprop-2-enoyl]amino}-2-hydroxybenzoic acid (C1), a new 5-aminosalicylic acid (5-ASA) derivative, has proven to be an antioxidant in vitro and an anti-inflammatory agent in mice. The in vivo inhibition of myeloperoxidase was comparable to that of indomethacin. The aim of this study was to take another step in the preclinical evaluation of C1 by examining acute toxicity with the up-and-down OECD method and pharmacokinetic profiles by administration of the compound to Wistar rats through intravenous (i.v.), oral (p.o.), and intraperitoneal (i.p.) routes. According to the Globally Harmonized System, C1 belongs to categories 4 and 5 for the i.p. and p.o. routes, respectively. An RP-HPLC method for C1 quantification in plasma was successfully validated. Regarding the pharmacokinetic profile, the elimination half-life was approximately 0.9 h with a clearance of 24 mL/min after i.v. administration of C1 (50 mg/kg). After p.o. administration (50 mg/kg), the maximum plasma concentration was reached at 33 min, the oral bioavailability was about 77%, and the compound was amply distributed to all tissues evaluated. Therefore, C1 administered p.o. in rats is suitable for reaching the colon where it can exert its effect, suggesting an important advantage over 5-ASA and indomethacin in treating ulcerative colitis and Crohn's disease.