Rapid Screening of Chemical Components in Salvia miltiorrhiza with the Potential to Inhibit Skin Inflammation.

Hyaluronidase possesses the capacity to degrade high-molecular-weight hyaluronic acid into smaller fragments, subsequently initiating a cascade of inflammatory responses and activating dendritic cells. In cases of bacterial infections, substantial quantities of HAase are generated, potentially leading to severe conditions such as cellulitis. Inhibiting hyaluronidase activity may offer anti-inflammatory benefits. Salvia miltiorrhiza Bunge, a traditional Chinese medicine, has anti-inflammatory properties. However, its effects on skin inflammation are not well understood. This study screened and evaluated the active components of S. miltiorrhiza that inhibit skin inflammation, using ligand fishing, enzyme activity assays, drug combination analysis, and molecular docking. By combining magnetic nanomaterials with hyaluronidase functional groups, we immobilized hyaluronidase on magnetic nanomaterials for the first time in the literature. We then utilized an immobilized enzyme to specifically adsorb the ligand; two ligands were identified as salvianolic acid B and rosmarinic acid by HPLC analysis after desorption of the dangling ligands, to complete the rapid screening of potential anti-inflammatory active ingredients in S. miltiorrhiza roots. The median-effect equation and combination index results indicated that their synergistic inhibition of hyaluronidase at a fixed 3:2 ratio was enhanced with increasing concentrations. Kinetic studies revealed that they acted as mixed-type inhibitors of hyaluronidase. Salvianolic acid B had Ki and Kis values of 0.22 and 0.96 µM, respectively, while rosmarinic acid had values of 0.54 and 4.60 µM. Molecular docking revealed that salvianolic acid B had a higher affinity for hyaluronidase than rosmarinic acid. In addition, we observed that a 3:2 combination of SAB and RA significantly decreased the secretion of TNF-α, IL-1, and IL-6 inflammatory cytokines in UVB-irradiated HaCaT cells. These findings identify salvianolic acid B and rosmarinic acid as key components with the potential to inhibit skin inflammation, as found in S. miltiorrhiza. This research is significant for developing skin inflammation treatments. It demonstrates the effectiveness and broad applicability of the magnetic nanoparticle-based ligand fishing approach for screening enzyme inhibitors derived from herbal extracts.

Structural basis of rosmarinic acid inhibitory mechanism on SARS-CoV-2 main protease.

The SARS-CoV-2 coronavirus is characterized by high mutation rates and significant infectivity, posing ongoing challenges for therapeutic intervention. To address potential challenges in the future, the continued development of effective drugs targeting SARS-CoV-2 remains an important task for the scientific as well as the pharmaceutical community. The main protease (Mpro) of SARS-CoV-2 is an ideal therapeutic target for COVID-19 drug development, leading to the introduction of various inhibitors, both covalent and non-covalent, each characterized by unique mechanisms of action and possessing inherent strengths and limitations. Natural products, being compounds naturally present in the environment, offer advantages such as low toxicity and diverse activities, presenting a viable source for antiviral drug development. Here, we identified a natural compound, rosmarinic acid, which exhibits significant inhibitory effects on the Mpro of the SARS-CoV-2. Through detailed structural biology analysis, we elucidated the precise crystal structure of the complex formed between rosmarinic acid and SARS-CoV-2 Mpro, revealing the molecular basis of its inhibitory mechanism. These findings not only enhance our understanding of the antiviral action of rosmarinic acid, but also provide valuable structural information and mechanistic insights for the further development of therapeutic strategies against SARS-CoV-2.

Anti-Obesity Activities of the Compounds from Perilla frutescens var. acuta and Chemical Profiling of the Extract.

Perilla frutescens var. acuta (Lamiaceae) is widely used not only as an oil or a spice, but also as a traditional medicine to treat colds, coughs, fever, and indigestion. As an ongoing effort, luteolin-7-O-diglucuronide (1), apigenin-7-O-diglucuronide (2), and rosmarinic acid (3) isolated from P. frutescens var. acuta were investigated for their anti-adipogenic and thermogenic activities in 3T3-L1 cells. Compound 1 exhibited a strong inhibition against adipocyte differentiation by suppressing the expression of Pparg and Cebpa over 52.0% and 45.0%, respectively. Moreover, 2 inhibited the expression of those genes in a dose-dependent manner [Pparg: 41.7% (5 µM), 62.0% (10 µM), and 81.6% (50 µM); Cebpa: 13.8% (5 µM), 18.4% (10 µM), and 37.2% (50 µM)]. On the other hand, the P. frutescens var. acuta water extract showed moderate thermogenic activities. Compounds 1 and 3 also induced thermogenesis in a dose-dependent manner by stimulating the mRNA expressions of Ucp1, Pgc1a, and Prdm16. Moreover, an LC-MS/MS chromatogram of the extract was acquired using UHPLC-MS2 and it was analyzed by feature-based molecular networking (FBMN) and the Progenesis QI software (version 3.0). The chemical profiling of the extract demonstrated that flavonoids and their glycoside derivatives, including those isolated earlier as well as rosmarinic acid, are present in P. frutescens var. acuta.

Database-aided ultrahigh-performance liquid chromatography Q-Exactive-Orbitrap tandem mass spectrometry putatively identifies 16 unexpected compounds and three anticounterfeiting pharmacopoeia quality markers for Perillae Fructus.

RATIONALE: Perillae Fructus (PF) is a common traditional Chinese medicine (TCM) for the treatment of asthma. It has not been effectively characterized by rosmarinic acid (RosA), which is currently designed as the sole quality indicator in the Chinese Pharmacopoeia. METHODS: This study introduced a database-aided ultrahigh-performance liquid chromatography equipped with quadrupole-Exactive-Orbitrap mass spectrometry (UHPLC/Q-Exactive-Orbitrap MS/MS) technology to putatively identify the compounds in PF, followed by literature research, quantum chemical calculation, and molecular docking to screen potential quality markers (Q-markers) of PF. RESULTS: A total of 27 compounds were putatively identified, 16 of which had not been previously found from PF. In particular, matrine, scopolamine, and RosA showed relatively high levels of content, stability, and drug-likeness. They exhibited interactions with the asthma-related target and demonstrated the TCM properties of PF. CONCLUSIONS: The database-aided UHPLC/Q-Exactive-Orbitrap MS/MS can identify at least 27 compounds in PF. Of these, 16 compounds are unexpected, and three compounds (matrine, scopolamine, and RosA) should be considered anticounterfeiting pharmacopoeia Q-markers of PF.

Protective Effect of Perilla Seed Meal and Perilla Seed Extract against Dextran Sulfate Sodium-Induced Ulcerative Colitis through Suppressing Inflammatory Cytokines in Mice.

An excessive inflammatory response of the gastrointestinal tract is recognized as one of the major contributors to ulcerative colitis (UC). Despite this, effective preventive approaches for UC remain limited. Rosmarinic acid (RA), an enriched fraction from Perilla frutescens, has been shown to exert beneficial effects on disease-related inflammatory disorders. However, RA-enriched perilla seed meal (RAPSM) and perilla seed (RAPS) extracts have not been investigated in dextran sulfate sodium (DSS)-induced UC in mice. RAPSM and RAPS were extracted using the solvent-partitioning method and analyzed with high-pressure liquid chromatography (HPLC). Mice with UC induced using 2.5% DSS for 7 days were pretreated with RAPSM and RAPS (50, 250, 500 mg/kg). Then, the clinical manifestation, colonic histopathology, and serum proinflammatory cytokines were determined. Indeed, DSS-induced UC mice exhibited colonic pathological defects including an impaired colon structure, colon length shortening, and increased serum proinflammatory cytokines. However, RAPSM and RAPS had a protective effect at all doses by attenuating colonic pathology in DSS-induced UC mice, potentially through the suppression of proinflammatory cytokines. Concentrations of 50 mg/kg of RAPSM and RAPS were sufficient to achieve a beneficial effect in UC mice. This suggests that RAPSM and RAPS have a preventive effect against DSS-induced UC, potentially through alleviating inflammatory responses and relieving severe inflammation in the colon.

Green Extraction of Depsidones and Depsides from Hypogymnia physodes (L.) Nyl. Using Natural Deep Eutectic Solvents.

Various studies have shown that Hypogymnia physodes are a source of many biologically active compounds, including lichen acids. These lichen-specific compounds are characterized by antioxidant, antiproliferative, and antimicrobial properties, and they can be used in the cosmetic and pharmaceutical industries. The main aim of this study was to optimize the composition of natural deep eutectic solvents based on proline or betaine and lactic acid for the extraction of metabolites from H. physodes. The design of the experimental method and the response surface approach allowed the optimization of the extraction process of specific lichen metabolites. Based on preliminary research, a multivariate model of the experiment was developed. For optimization, the following parameters were employed in the experiment to confirm the model: a proline/lactic acid/water molar ratio of 1:2:2. Such a mixture allowed the efficient extraction of three depsidones (i.e., physodic acid, physodalic acid, 3-hydroyphysodic acid) and one depside (i.e., atranorin). The developed composition of the solvent mixtures ensured good efficiency when extracting the metabolites from the thallus of H. physodes with high antioxidant properties.

Lipophilized rosmarinic acid: Impact of alkyl type and food matrix on antioxidant activity, and optimized enzymatic production.

In this study, the initial focus was on exploring the simultaneous impact of the oil-based food matrix and the polarity of rosmarinic acid derivatives on the antioxidant properties. Rosmarinic acid (RA) showed remarkable DPPH, FRAP, and ABTS radical scavenging activities, followed by methyl rosmarinate (MR) and ethyl rosmarinate (ER). In bulk oil, both conjugated dienes and p-AnV values reached a peak in the following order after 30 days: ER > MR > RA = BHT > control (no antioxidant). In the oil structured using monoacylglycerol, MR was more effective than ER and RA. For ethyl cellulose oleogel, emulsion, and gelled emulsion systems, RA was more effective. Additionally, after confirming the importance of the food matrix on the antioxidant activity of RA derivatives, the lipophilization of RA with ethanol was optimized as a model with Lipozyme 435 in hexane. A conversion yield of as high as 85.59% for ER was achieved, as quantified by HPLC-UV and confirmed by HPLC-DAD-ESI-qTOFMS.

Effects of the dose of administration, co-antioxidants, food matrix, and digestion-related factors on the in vitro bioaccessibility of rosmarinic acid - A model study.

This study aimed to determine the effect of the administration dose, combinations with co-antioxidants (vitamin C, caffeic acid, chlorogenic acid, catechin, rutin), and different food matrices (cooked and lyophilized hen eggs, chicken breast, soybean seeds, potatoes) on the potential bioaccessibility of rosmarinic acid (RA) in simulated digestion conditions, depending on the digestion stage (gastric and intestinal) and the contribution of physicochemical and biochemical digestion factors. The in vitro bioaccessibility of RA depended on the digestion stage and conditions. The physicochemical factors were mainly responsible for the bioaccessibility of RA applied alone. The higher RA doses improved its bioaccessibility, especially at the intestinal stage of digestion. Furthermore, the addition of vitamin C and protein-rich food matrices resulted in enhanced intestinal bioaccessibility of RA. In the future, the knowledge of factors influencing the bioaccessibility of RA can help enhance its favorable biological effects and therapeutic potential.

Bio-Guided Isolation of Alpha-Glucosidase Inhibitory Compounds from Vietnamese Lichen Roccella Montagnei.

A bio-guided isolation was applied to the Vietnamese lichen Roccella montagnei based on alpha-glucosidase inhibition. Six compounds were isolated and structurally elucidated, including a new ortho depside, montagneside A (1), together with five known compounds, sekikaic acid (2), lanost-7-en-3ß-ol (3), ethyl orsellinate (4), D-montagnetol (5), and D-erythrin (6). Their chemical structures were identified by extensive 1D and 2D NMR analysis, high-resolution mass spectroscopy, and comparisons with those reported in the literature. D-Erythrin (6), a major component, was selected for further modification using Smiles rearrangement. Three erythritol derivatives 6a-6c were synthesized. Compounds 1-3, 6, and 6a-6c were evaluated for alpha-glucosidase inhibition. Compounds 2 and 6a-6c showed significant alpha-glucosidase inhibition with IC50 values ranging from 7.9 to 149 µM, respectively. Molecular docking was applied to the most active compound 6a to clarify the inhibitory mechanism.

α-Glucosidase Inhibitory Components from Garcinia pedunculata Fruits.

From Garcinia pedunculata Roxb. fruits, two undescribed aromatic compounds including a benzofuran and a depsidone derivative, and a new natural product, together with four known compounds were isolated. Through the analysis of spectroscopic data, high resolution mass spectrum and calculated nuclear magnetic resonance, their structures were determined. The α-glucosidase inhibitory activity of the isolates was evaluated. And compound 3 exhibited a moderate inhibitory effect on α-glucosidase. The molecular docking of compound 3 was performed to elucidate the interaction with α-glucosidase.

Mechanism Behind the Programmed Biosynthesis of Heterotrimeric Fungal Depside Thielavin A.

Thielavin A (1) is a fungal depside composed of one 3-methylorsellinic acid and two 3,5-dimethylorsellinic acid units. It displays diverse biological activities. However, the mechanism underlying the assembly of the heterotrimeric structure of 1 remains to be clarified. In this study, we identified the polyketide synthase (PKS) involved in the biosynthesis of 1. This PKS, designated as ThiA, possesses an unusual domain organization with the C-methyltransferase (MT) domain situated at the C-terminus following the thioesterase (TE) domain. Our findings indicated that the TE domain is solely responsible for two rounds of ester bond formation, along with subsequent chain hydrolysis. We identified a plausible mechanism for TE-catalyzed reactions and obtained insights into how a single PKS can selectively yield a specific heterotrimeric product. In particular, the tandem acyl carrier protein domains of ThiA are critical for programmed methylation by the MT domain. Overall, this study highlighted the occurrence of highly optimized domain-domain communication within ThiA for the selective synthesis of 1, which can advance our understanding of the programming rules of fungal PKSs.

A New Rare Halogenated Depside from Lichen and Study of its Anti-Proliferative Activity.

Lichens are a symbiotic association of algae and fungus, belonging to the family Parmeliaceae. Some lichen species are edible and used as an active ingredient for preparation of exotic spices as well as folklore medicine to cure different kinds of ailments. A specimen of lichen was collected from Munner in the Kerala State of South India for chemical profiling. Chemical analyses of the diethyl ether extract of the defatted lichen led to the isolation of six phenols 1-6 with variation of relative abundance. Amongst them, the relative abundance of compound 3 was the greatest (1 % of crude extract) and it was identified as atranorin. The structures of known compounds were confirmed by comparison of their 1H-NMR, 13C NMR, and mass data with published values available in the literature. In vitro bioassay for anti-proliferative activity of these compounds has been conducted against various human cancer cell lines in comparison with paclitaxel as control using SRB assay. Interestingly, a new compound 5 was found along with previously reported compounds from this lichen. This new compound was designated as fluoroatranorin 5 which was reported for the first time herein. The structural characterization of a new depside was determined by spectral methods such as 1H-NMR, 13C NMR, 19F NMR, IR, LC-HRESI-MS, and LC-MS/MS study. Its structure was confirmed by single crystal X-ray diffraction study. This new compound was designated as fluoroatranorin 5 which was reported first time herein. Anti-proliferative activity of all these compounds was evaluated against six different cancer cell lines. The inhibitory activity, IC50 value of compounds 1-3 and 5 exhibited at 99.64, 102.04, 109.20, 53.0 and 2.4 µM on cancer cell lines HT-29 (colon), Hela (cervical), HT-29, HPAC (pancreas) and A2780 (ovarian cancer cell line) respectively in comparison with paclitaxel as control. The new compound 5 exhibited significant activity with IC50 value 2.4 µM on A2780 ovarian cancer cell line.

Complexation of bovine lactoferrin with selected phenolic acids via noncovalent interactions: Binding mechanism and altered functionality.

Noncovalent interactions of 4 selected phenolic acids, including gallic acid (GA), caffeic acid (CA), chlorogenic acid (CGA), and rosmarinic acid (RA) with lactoferrin (LF) were investigated. Compound combined with LF in the binding constant of CA > GA > RA > CGA, driven by van der Waals and hydrogen bonding for GA, and hydrophobic forces for others. Conformation of LF was affected at secondary and ternary structure levels. Molecular docking indicated that GA and CA located in the same site near the iron of the C-lobe, whereas RA and CGA bound to the C2 and N-lobe, respectively. Significantly enhanced antioxidant activity of complexes was found compared with pure LF, as demonstrated by 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azinobis(2-ethylbenzothiazoline-6-sulfonate) (ABTS), and ferric reducing antioxidant power (FRAP) models. Caffeic acid, CGA, and RA significantly decreased the emulsifying stability index and improved foam ability of LF, and the effect of CA and RA was the most remarkable, respectively.

Anti-Inflammatory and Antioxidant Effects of Rosmarinic Acid Trimetallic (Cu0.5Zn0.5Fe2O4) Nanoparticles.

Newly, green metallic-nanoparticles (NPs) have received scientists' interest due to their wide variable medicinal applications owned to their economical synthesis and biologically compatible nature. In this study, we used rosmarinic acid (RosA) to prepare Cu0.5Zn0.5FeO4 NPs and later encapsulated them using PEG polymer. Characterization of NPs was done using the XRD method and SEM imaging. Further, we explored the encapsulated NPs for anti-inflammatory properties by downregulating the expression of pro-inflammatory cytokines mRNA in LPS-stimulated Raw 264.7 cells. Besides, employing DPPH, NO and ABTS radical scavenging assays to examine the antioxidant activity of the synthesized Cu0.5Zn0.5FeO4 NPs. Cu0.5Zn0.5FeO4 NPs revealed moderate antioxidant activity by scavenging DPPH and nitric oxide. We demonstrated that the NPs showed high potential anti-inflammatory activity by suppressing the mRNA and protein levels of pro-inflammatory cytokines in a dose-dependent manner, in LPS-induced Raw 264.7 cells. To our best knowledge, this is the first report where RosA was found to be a suitable phyto source for the green synthesis of Cu0.5Zn0.5FeO4 NPs and their in vitro anti-inflammatory and antioxidant effects. Taken together, our findings suggest that the RosA is a green resource for the eco-friendly synthesis of Cu0.5Zn0.5FeO4/PEG NPs, which further can be employed as a novel anti-inflammatory therapeutic agent.

Botryorhodines K and L, two new cytotoxic depsidones from a fungus of the genus Arcopilus.

Botryorhodines K (1) and L (2), two new depsidone derivatives, along with one known metabolite, 4-O-demethylbarbatic acid (3), were isolated from the culture extract of a fungus of the genus Arcopilus. The structures of 1‒3 were determined by the analysis of NMR and MS spectral data and the absolute configuration of 1 was established by single-crystal X-ray diffraction analysis. Compounds 1 and 2 showed antimicrobial activity against Gram-positive bacteria and cytotoxicity against murine leukemia P388 cells.

The Influence of Methyl Jasmonate on Expression Patterns of Rosmarinic Acid Biosynthesis Genes, and Phenolic Compounds in Different Species of Salvia subg. Perovskia Kar L.

Salvia yangii B.T. Drew and Salvia abrotanoides Kar are two important fragrant and medicinal plants that belong to the subgenus Perovskia. These plants have therapeutic benefits due to their high rosmarinic acid (RA) content. However, the molecular mechanisms behind RA generation in two species of Salvia plants are still poorly understood. As a first report, the objectives of the present research were to determine the effects of methyl jasmonate (MeJA) on the rosmarinic acid (RA), total flavonoid and phenolic contents (TFC and TPC), and changes in the expression of key genes involved in their biosynthesis (phenylalanine ammonia lyase (PAL), 4-coumarate-CoA ligase (4CL), and rosmarinic acid synthase (RAS)). The results of High-performance liquid chromatography (HPLC) analysis indicated that MeJA significantly increased RA content in S. yungii and S. abrotanoides species (to 82 and 67 mg/g DW, respectively) by 1.66- and 1.54-fold compared with untreated plants. After 24 h, leaves of Salvia yangii and Salvia abrotanoides species treated with 150 M MeJA had the greatest TPC and TFC (80 and 42 mg TAE/g DW, and 28.11 and 15.14 mg QUE/g DW, respectively), which was in line with the patterns of gene expression investigated. Our findings showed that MeJA dosages considerably enhanced the RA, TPC, and TFC contents in both species compared with the control treatment. Since increased numbers of transcripts for PAL, 4CL, and RAS were also detected, the effects of MeJA are probably caused by the activation of genes involved in the phenylpropanoid pathway.

Deciphering the therapeutical potentials of rosmarinic acid.

Lemon balm is herbal tea used for soothing stomach cramps, indigestion, and nausea. Rosmarinic acid (RA) is one of its chemical constituents known for its therapeutic potentials against cancer, inflammatory and neuronal diseases such as the treatment of neurofibromatosis or prevention from Alzheimer's diseases (AD). Despite efforts, recovery and purification of RA in high yields has not been entirely successful. Here, we report its aqueous extraction with optimal conditions and decipher the structure by nuclear magnetic resonance (NMR) spectroscopy. Using various physical-chemical and biological assays, we highlight its anti-aggregation inhibition potentials against the formation of Tau filaments, one of the hallmarks of AD. We then examine its anti-cancer potentials through reduction of the mitochondrial reductase activity in tumor cells and investigate its electrochemical properties by cyclic voltammetry. Our data demonstrates that RA is a prominent biologically active natural product with therapeutic potentials for drug discovery in AD, cancer therapy and inflammatory diseases.

A Comprehensive Review of Rosmarinic Acid: From Phytochemistry to Pharmacology and Its New Insight.

Polyphenolic acids are the widely occurring natural products in almost each herbal plant, among which rosmarinic acid (RA, C18H16O8) is well-known, and is present in over 160 species belonging to many families, especially the Lamiaceae. Aside from this herbal ingredient, dozens of its natural derivatives have also been isolated and characterized from many natural plants. In recent years, with the increasing focus on the natural products as alternative treatments, a large number of pharmacological studies have been carried out to demonstrate the various biological activities of RA such as anti-inflammation, anti-oxidation, anti-diabetes, anti-virus, anti-tumor, neuroprotection, hepatoprotection, etc. In addition, investigations concerning its biosynthesis, extraction, analysis, clinical applications, and pharmacokinetics have also been performed. Although many achievements have been made in various research aspects, there still exist some problems or issues to be answered, especially its toxicity and bioavailability. Thus, we hope that in the case of natural products, the present review can not only provide a comprehensive understanding on RA covering its miscellaneous research fields, but also highlight some of the present issues and future perspectives worth investigating later, in order to help us utilize this polyphenolic acid more efficiently, widely, and safely.

Biomedical features and therapeutic potential of rosmarinic acid.

For decades, the use of secondary metabolites of various herbs has been an attractive strategy in combating human diseases. Rosmarinic acid (RA) is a bioactive phenolic compound commonly found in plants of Lamiaceae and Boraginaceae families. RA is biosynthesized using amino acids tyrosine and phenylalanine via enzyme-catalyzed reactions. However, the chemical synthesis of RA involves an esterification reaction between caffeic acid and 3,4-dihydroxy phenyl lactic acid contributing two phenolic rings to the structure of RA. Several studies have ascertained multiple therapeutic benefits of RA in various diseases, including cancer, diabetes, inflammatory disorders, neurodegenerative disorders, and liver diseases. Many previous scientific papers indicate that RA can be used as an anti-plasmodic, anti-viral and anti-bacterial drug. In addition, due to its high anti-oxidant capacity, this natural polyphenol has recently gained attention for its possible application as a nutraceutical compound in the food industry. Here we provide state-of-the-art, flexible therapeutic potential and biomedical features of RA, its implications and multiple uses. Along with various valuable applications in safeguarding human health, this review further summarizes the therapeutic advantages of RA in various human diseases, including cancer, diabetes, neurodegenerative diseases. Furthermore, the challenges associated with the clinical applicability of RA have also been discussed.

Intramolecular transesterification of depsides yields fluorescent 1H-isochromen-1-ones: Application as a chemical probe for lichen determination.

The reactivity of eight purified depsides obtained from six european lichens and that display as 2-oxoalkyl chain in ortho-position of the ester bond was explored. These depsides were found to lead to 1H-Isochromen-1-ones, which exhibit a distinctive blue fluorescence at 365 nm, in the presence of a 10% aqueous solution of KOH. A mechanistic explanation, involving the formation of an enolate intermediate and intramolecular transesterification, was proposed and validated by DFT. By exploiting this fluorescent phenomenon, we conceived a chemical probe (the KUV probe) that is useful for lichen determination, as exemplified on a selection of European Porpidia species.